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aa Raja
Section 1hemistry Investigation
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Table of ContentsIntroduction........................................................................................................... 3
General information........................................................................................3
Natural processes and manufacture................................................................3
ses.................................................................................................................3
Collision theory......................................................................................................!
"ressure................................................................................................................. !
Surface #rea..........................................................................................................!
Catalysts................................................................................................................ $
General info..................................................................................................... $
%ypes of catalysts............................................................................................$
&n'ymes (catalase)................................................................................................*
General function..............................................................................................*
Structure of catalase....................................................................................... *
+unction of catalase........................................................................................ *
"ossi,le -echanisms.......................................................................................
-echanism of catalase /ith 0o.....................................................................
Inhi,ition......................................................................................................... 2
p0.......................................................................................................................... 2
%emperature..........................................................................................................
#rrhenius &4uation..........................................................................................
&5ample........................................................................................................ 16
Concentration...................................................................................................... 11
General..........................................................................................................11
-ichaelis7-enten 8inetics............................................................................. 11
9ine/eaver7:ur; "lot.....................................................................................1
Rate e4uations.....................................................................................................13
#ims..................................................................................................................... 1!
-ethods of ma;ing solutions...............................................................................1$
0ydrogen "ero5ide solution...........................................................................1$
Catalase solution...........................................................................................1*
"otassium (I<) -anganate.............................................................................1
-ethods of measuring rate..................................................................................12
Gas syringe................................................................................................... 1
"age = 1
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Su,a seal gas syringe....................................................................................6
Inverted ,urette............................................................................................1
......................................................................................................................
%itration.........................................................................................................3
Ris; #ssessment..................................................................................................$
:i,liography.........................................................................................................31
Introduction...................................................................................................31
9ist of +igures................................................................................................ 3
9ist of &4uations............................................................................................33
-ethods.........................................................................................................33
Ris; #ssessment............................................................................................3!
"age =
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IntroductionGeneral aims> I /ill investigate the ;inetics of the decomposition of hydrogen
pero5ide /ith the en'yme catalase e5perimentally ,y varying the concentration
and temperature.
General information
%he e4uation for the decomposition of hydrogen pero5ide /ith catalase>
catalase0?(a4) 0?(l) @ ?(g) equation 1
#s illustrated in Agure 1B hydrogen pero5ide is a
compound of hydrogen and o5ygen (it can ,e thought of
as /ater /ith an e5tra o5ygen atom). "ure anhydrous
hydrogen pero5ide is a colourless li4uid that rapidly
decomposes into o5ygen and /ater. %he ,ond ,et/een the
t/o o5ygen atoms of the molecule is very /ea;B so the
molecule can very easily ,rea; do/nB ma;ing it reactive
and a po/erful o5idising agent. 1
Natural processes and manufacture
%he human ,ody produces hydrogen pero5ide in the
immune system. It is produced naturally as ,y7product of o5ygen meta,olism.
-ost living ,eings have pero5idases en'ymes that decompose small amounts of
hydrogen pero5ide present into /ater and o5ygen to prevent tissue damage,ecause it poisonous. 0ydrogen "ero5ide can
,e produced indirectly ,y hydrogenation and
o5idation of ananthra4uinone. # direct
method of hydrogenation not only
hydrogenates ? in 0 ? ,ut also converts
0? into /ater.
Uses
0ydrogen pero5ide is used to treat /aste /ater and se/age from industrial and
domestic sources and for deto5ifying organic pollutants in the environment. It isalso used as a commercial disinfectant and antimicro,ial agent. Since it is
e5tremely reactive it is used as a component in some types of roc;et fuel. #s a
disinfectantB hydrogen pero5ide has ,een sho/n to ,e generally eEective /ith
very safe ,y7products. 3 #t lo/ concentrations (around 37*F) it is used in
pero5ide7,ased hair dyes. !
%he rate of decomposition is dependent on the temperature and concentration of
the pero5ideB as /ell as the presence of impurities and sta,ili'ers. %he use of a
catalyst (such as manganese dio5ideB silverB or the en'yme catalase) increases
the rate of decomposition. $
"age = 3
Figure 1
Shape of hydrogen peroxide
Figure
!roduction of "ydrogen !eroxide by#nthraquinone !rocess
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Collision theory
-olecules need to collide for a reaction to ta;eplace. :ut not all collisions /ill result in a reaction.
+igure 3 demonstrates the orientation in /hich the
molecules must ,e in order for them to react for
that particular reaction. * #ctivation energy is the
energy the reactants need to overcome the
repulsive electron forces ,et/een approaching
molecules and to ,rea; the e5isting ,onds in the
reacting molecules.
# decomposition is an e5ample of a unimolecular
reaction. %his is /here a single reactant moleculetransforms into one or more products. %he rate at /hich a su,stance
decomposes depends on its concentration and it is
often Arst order. %he reaction I am investigating is
e5othermic as heat is released and the reactants have more energy than the
products as sho/n in Agure !.
%he rate of a chemical reaction can ,e changed ,y>
• the pressure of the system (if some
reacting molecules are a gas)• p0
• surface area (if some reactants are in solidphase)
• adding a catalyst
• the temperature
• the concentration of catalase (catalyst) andhydrogen pero5ide (su,strate)
!ressureIncreasing the pressure on a reaction increases the rate of
reaction only /hen the reactants are gaseous. "ressure is
usually increased ,y reducing the volume. If pressure is
increased then more particles are present in the same volume and there /ill ,e a
higher collision fre4uency leading to an increased rate of reaction. I /ill not need
to ta;e the eEect of pressure into account as the reactants are not gases ,ut I
could represent the rate as the change in pressure as o5ygen is produced.
Surface #rea
Increasing the surface area ,y cutting it into smaller pieces or grinding it into apo/der /ill increase the rate of reaction. %his is ,ecause more particles are
"age = !
Figure $
Figure %
The higher the &a of a reactionthe smaller the amount of
energetic collisions present andthe slo'er the reaction
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e5posed to reactantB therefore there are more fre4uent collisions and rate
increases. Surfaces area is not something I need to consider in my e5periments
as I /ill ,e /or;ing /ith li4uids.
Catalysts
General info
Catalysts are compounds that increase the reaction rate ,y providing an
alternative path/ay for the reaction /ith lo/er activation energy.
%hey /ill have
no eEect on the energetic characteristics of the hydrogen pero5ide and catalase
and the ,arriers ,et/een them.
%he amount of a catalyst does not change during a reactionB it is not consumed
as part of the reaction process. Catalysts do not aEect the e4uili,rium state.
%his is ,ecause it speeds up ,oth the for/ard and ,ac;/ard reaction ,y the
same amount. If ,oth reaction rates are increased to the same e5tentB only the
rate at /hich e4uili,rium is reached is increased.
Types of catalysts
Catalysts can ,e categori'ed into t/o types> homogeneous and heterogeneous.
0omogeneous catalysts are those /hich e5ist in the same phase as the
reactantsB /hile heterogeneous catalysts are not in the same phase as the
reactants.
NormallyB heterogeneous catalysis involves the use of solid catalysts placed in a
li4uid reaction mi5ture. 0omogeneous catalysis means more eEective mi5ing of the catalyst /ith the reaction mi5ture heterogeneous catalysis allo/s recovery
of e5pensive catalystsB /hich is useful for manufacturing processes. 2
"age = $
Figure (Figure )0eterogeneous
• %he reactants are adsor,ed on tothe surface of the catalyst. %his isa chemical reaction as there is aninteraction ,et/een the electronsof the reactants and the atoms onthe catalyst
• %he adsor,ed reactants are free tomigrate over the surface of thecatalyst
•
Hhen the reactants meet they arefree to react ,ut still attached tothe surface
• +inally the products of the reactionare desor,ed from the surfaceallo/ing them to move a/ay andfreeing up area for more reactions
0omogeneous
# t/o stage reaction proAle for a
catalytic cycle (&a activation energy)
• &a1>activation energy thatJs leads
to an intermediate comple5
forming.
• &a>activation energy for the
changing of the intermediate into
products.
• &a3>activation energy of the
uncatalysed reaction.
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&n*ymes +catalase,
General function
Catalase is an e5ample of a ,iological catalyst it increases the rate of reaction
/ithout ,eing changed or depleted itself. It is a protein it has a very precise
three7dimensional shapeB /hich forms one speciAc active site on the en'yme
/here molecules. &ach en'yme can only convert one ;ind of su,strate molecule
into one ;ind of product molecules.
%he ,asic mechanism ,y /hich catalase catalyse the reaction ,egins /ith the,inding of the hydrogen pero5ide to the active site on catalase. %he active site isthe speciAc region of the catalase /hich com,ines /ith the 0? molecule. %he,inding of hydrogen pero5ide to catalase /ill cause changes in the distri,ution of electrons in the chemical ,onds of the catalase and cause the reaction that leadsto the formation of o5ygen and /ater. %hese are released from the catalasesurface to regenerate the en'yme for another reaction cycle.
%he en'yme is the loc; and so hydrogen pero5ideis the ;ey. ?nly the correct ;eyKsu,strate Ats intothe ;eyhole or the loc;Kactive site. %he activesite of the en'yme has a uni4ue shape that iscomplementary to the shape of the hydrogen pero5idemolecule. &n'ymes speciAcally react /ith only one or avery fe/ similar compounds as sho/n ,y the loc; and;ey model in Agure .
Structure of catalaseCatalase is a tetramer (a protein /ith four su,units) of four polypeptide chainsB
each over $66 amino acids long 11 as sho/n in Agure 2.
"age = *
Figure -
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Function of catalase
%he catalyst I /ill ,e using for this investigation
is catalase /hich is a homogeneous catalyst. It is
made naturally in almost all living organisms and
are important to life as it helps the ,ody ,rea;
do/n hydrogen pero5ide (a po/erful and harmfulo5idi'ing agent) and prevents the accumulation
of car,on dio5ide ,u,,les in the ,lood. Catalase
is very eEective one molecule of catalase can
decompose millions of hydrogen pero5ide
molecules. It also uses hydrogen pero5ide to
o5idi'e potentially harmful to5ins li;e
formaldehydeB formic acidB alcoholB and phenol 16 from occurring in the ,ody.
%he optimum p0 for catalase is roughly as this usually the p0 of the cells
/here catalase carries out its functions. Hhile p0 can vary ,y speciesB the
catalase I /ill ,e using is derived from ,ovine liver /hich is also roughly neutral.
!ossible .echanisms
&n'ymes can catalyse reactions through a range of mechanisms. Some of these
include>
• :ond strain> induced structural rearrangement that ta;e place /ith the
,inding of su,strate and en'yme produce strained su,strate ,onds that
can easily reach the transition state
• "ro5imity and orientation> causes the en'yme7su,strate interaction to
orient reactive groups in a /ay that ,ring them into close pro5imity /ith
one another• "roton donors and acceptors> the presence of acidic or ,asic groups can
aEect ,ond polari'ation and reaction speed (the acid is often a donor
/hereas the ,ase is an acceptor)• &lectrostatic catalysis> side chains or cofactors (non7protein part) can
sta,ili'e the charge on the transition state
• Covalent catalysis> the su,strate is oriented to the active site in such a
/ay that a covalent intermediate forms ,et/een the en'yme and
su,strate. %he covalent comple5 is more reactive than the su,strate itself
originally /as. 13
.echanism of catalase 'ith "o
%he protein has a precise 3D structure /hen it is activeB /hich contains a
channel into /hich the hydrogen pero5ide can diEuse. In the channel is a hemegroup /hich is an iron molecule ,ound to the centre of a ring7li;e structure called
"age =
Figure /
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a porphyrin ring. %he heme group in catalase is important to the reactionB
,ecause it can ,e o5idi'ed from +e(III) to the less common +e(I<) form. %his is the
Arst part of the reaction>
0? @ +e(III) L0 ? @?+e(I<)equation
0ydrogen pero5ide has ,ound to the heme group and o5idi'ed it to +e(I<).
%he en'yme goes ,ac; to the +e (III) form ,y reducing the second molecule of
hydrogen pero5ide to /ater.
0? @?+e(I<)L0?@+e(III) equation $
%he highly7o5idi'ing +e(I<) reacts /ith the second pero5ide moleculeB releasing
/ater and an o5ygen molecule. 1
-ost en'yme reactions have optimum conditions due to irreversi,le damage
(denaturation) done ,y p0 and temperatures as sho/n ,y the graphs ,elo/>
%his means there is a limited range of temperatures I can use ,ecause thereaction may ,e too slo/ to measure at lo/er or higher temperatures. 9o/ertemperatures mean the molecules /ill not have enough ;inetic energy to resultin successful collisions and then reactions. 0o/everB approaching hightemperaturesB the reaction rate /ill start to decrease as the en'ymes denature
and cannot ,rea; do/n hydrogen pero5ide to produce o5ygen and /ater.SimilarlyB changes in the p0 can limit the reaction so I need to ensure the p0 is
"age = 2
Figure 0
Summary of e4uations and 3 to sho/ ho/ the
overall e4uation of 06 decomposition is
derived
Figure 1Figure 11
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;ept constant at the optimum value in my e5periment. #ny deviations around theoptimum /ill cause the catalase to denature and rate of reaction to decrease.
Inhibition
&n'yme inhi,itors can reduce or completely inhi,it the en'yme catalytic activity
either reversi,ly or permanently. Inhi,itors can ,loc; or modify the active site/hich means that the su,strate is prevented from ,eing ,ro;en do/n. %hose that
have similar structure to the su,strate and occupy the active site are called
competitive inhi,itors and those that attach to other parts of the en'yme
molecule and distort the shape (and therefore active site) are non7competitive.
+or catalaseB copper sulphate is a non7competitive inhi,itor. Cyanide is a
competitive inhi,itor ,ecause it ,inds to the active site and prevents an en'yme7
su,strate comple5 forming /ith catalase and hydrogen pero5ide. Since cyanide
is very to5icB another /ay the reaction can ,e stopped is ,y the use of
concentrated sulfuric acid /hich lo/ers the p0 so much that all of the catalase
molecules ,ecome denatured and further decomposition is halted. I /ill usecopper sulphateB sulphuric acid and ethanol 1! to stop the reaction at certain
points so the concentration of hydrogen pero5ide at that point can ,e measured.
p" %his is the measure of the acidity or concentration of hydrogen ions in a solution.
#s catalase is an en'yme (and therefore a protein made up of amino acids /ith
7N0 and 7C??0 groups on side chains) the ,alance of ?07 and 0@ ions inMuences
the shape of the active site so that it is most complementary to the shape of
hydrogen pero5ide. If the ioni'ation state of these amino acids is alteredB the
,onds that create the shape of the en'yme /ill ,e altered and the su,strate /ill
not ,ind to the active site and ,e catalysed. DiEerent en'ymes have diEerent
optimum p0. In this investigation I /ill not ,e measuring p0 ,ecause there /ill
,e too many varia,les to consider and there /ill ,e too many calculations.
0o/ever it can ,e ;ept constant using p0 ,uEer solution. # p0 ,uEer
Temperature#n increase in temperature causes a rise in the energy of the molecules involved
in the reaction. #s a result the rate of the reaction increases and conversely the
rate of reaction /ill decrease /ith a decrease in temperature.
"age =
Figure 1
Hhere temperature of % %1
Increasing the temperature /ill ma;e
the pea; move lo/er and to the right.
%he area under the curves should ,e
e4ual as the num,er of particles are
the same. Increasing the temperature/ill increase the rate as a greater
proportion of particles /ill have the
minimum activation energy.
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In en'yme controlled reactions /hen the temperature is too high the ,onds
(especially hydrogen ,onds) ,rea; and this causes the shape of the en'yme to
change. %his causes the active site to change shape and the en'yme can no
longer accept su,strate molecules. I cannot carry out the e5periments at higher
temperatures as the en'yme denatures the rate may ,e too small to measure.
#rrhenius &quation
%he #rrhenius e4uation can ,e used to sho/ the eEect of a change of
temperature on the rate constant and therefore the rate of the reaction.
%he #rrhenius e4uation can ,e /ritten in a non7e5ponential form /hich /ill ,emore convenient to use and interpret graphically. %his e4uation is of a straightline (ym5@c).
ln k =− Ea
R
1
T + ln A equation %
I /ill need to determine the rate constant (page 13 sho/s ho/) /hile changing
the temperature and plot a graph of
1
T against ln k . -easuring the gradient
"age = 16
• k is the rate constant
•
A is ;no/n as the pre7e5ponential factor.It is the fraction of molecules that /ouldreact if the activation energy /as a,ove6B or if the ;inetic energy of allmolecules are a,ove &a
• Ea is the activation energy
• %his is the minimum energy needed forthe reaction to occur. %o At this into the
Figure 1$
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/ill then give− Ea
R B /hich /ill give me the activation energy for the reaction.
1$
#lternativelyB the activation energy can ,e found alge,raically ,y using t/o rateconstants and t/o corresponding reaction temperatures ,ut it is less accurate.
lnk 1
k 2=− Ea
R ( 1T 1−
1
T 2 ) equation )
&xample
%he rate constant for the reaction 0(g) @ I(g)L 0I(g) is $.! 5 16 7! -71s71 at 3* oC.
#t !16 oC the rate constant /as found to ,e .2 5 167 - 71s71.
He can calculate the activation energy using the integrated form of the
#rrhenius e4uation as /e ;no/ the rate constant for the reaction at t/o diEerent
temperatures. 3*oC is $8 and !16oC is *238.
ln 5.4 x 10
−4mol
−1s−1
2.8 x 10−4mol
−1s−1=
− Ea
R ( 1599 K − 1
683 K )
−3.9484=− Ea
R (2.053 x10−4 K −1)
Ea=1.923 x 104 K x 8.314 J /mol k
Ea=1.60 x 105
J /mol
Concentration
General
-ore collisions ta;ing place means there /ill ,e more successful collisions and
this /ill lead to a faster reaction rate. Hhen more hydrogen pero5ide molecules
are addedB more en'yme7su,strate comple5es can ,e formed and the rate ofreaction increases. &ventuallyB increasing this concentration further /ill have no
"age = 11
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eEect ,ecause the active sites /ill ,e full and so no more en'yme7su,strate
comple5es can form. #s the catalase concentration increasesB there /ill ,e more
active sites availa,le and the reaction can happen faster. 0o/ever there /ill
come a point /here increasing its concentration ,eyond a certain point /ill have
no eEect as the hydrogen pero5ide concentration /ill ,ecome the limiting factor.
.ichaelis2.enten 3inetics
%he rate of the catalase reaction follo/s one of the general e4uations foren'yme7catalysed reactionsB the -ichaelis7-enten e4uation. %he rate of thecatalase reaction /ith hydrogen pero5ide is>
reactionrate V 0=
V max (concentration of hydrogen peroxide )
K m+ (concentration of hydrogen peroxide ) equation (
V 0 is the inital velocity of the reaction
V max is the ma5imum rate of the reactionK m, the -ichaelis7-enten constant is the concentration of the su,strate at /hich
the rate of the reaction is one half the <ma5. It can also ,e thought of as a
measure of ho/ /ell the su,strate comple5es /ith an en'yme (;no/n as its
,inding aPnity). #n e4uation /ith a lo/ 8 m value sho/s a large ,inding aPnityB
as the reaction /ill reach <ma5 4uic;er. 1*
8 m is also the concentration of the su,strate in mol dm 73 /hich produces a
reaction rate half of <ma5.
%he -ichaelis7-enten e4uation sho/s that the reaction rates for en'ymereactions saturate 7 they reach a ma5imum reaction rate (<ma5)B and stay at thatma5imum reaction rate regardless of ho/ much more hydrogen pero5ide isadded ,eyond that point.
4ine'ea5er26ur7 !lot
Simply graphing rate and su,strate concentration /ould not give <ma5 (therefore
8 m) as it is an asymptote. Its value can only ,e certain if the reaction ta;es place
"age = 1
Figure 1%
<ma5 is approached asymptotically as
concentration of su,strate increases.
It is the ma5imum rate at /hich an
en'yme catalyses a reaction and
e5pressed as the amount of productformed per minute.
# large 8m re4uires high su,strateconcentrations to achieve ma5imumreaction velocity. %his means theen'yme isnJt as eEective
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at an inAnite concentration of su,strate /hich is impossi,le to do to
e5perimentally.
%he 9ine/eaver7:ur; dou,le reciprocal plot rearranges the -ichaelis7-enten
e4uation into a linear form>
1
V =
K m
V max
. 1
[S]+
1
V maxequation -
8ate equations
%he order of an e4uation is /hat the concentration of a su,stance is raised to in
the rate e4uation. %he greater that num,erB the greater eEect it /ill have onrate. %he order of a reaction must ,e determined e5perimentally.
"age = 13
Figure 1)
"lotting 1Kv against 1KS gives a
straight line /here>
• y intercept is 1K <ma5
• gradient is 8m K <ma5
• 5 intercept is 71K 8m
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Chemical reactions are categori'ed on the ,asis of ;inetics as 'ero7orderB Arst7
orderB second7order>
• Qero7orderthere is no eEect on the initial rate of reaction r; (ms 71 )
• +irst7order the initial rate of reaction dou,les r=k[A] ( s71 )
• Second order the initial rate of reaction 4uadruples or r=[A][B] or r=k[A]2
( m71.s71 )
0o/ever these are the general units and /ill vary on /hat method is used to
measure # and :.
%he order of a reaction can ,e esta,lished from a concentration7time graph ,ut it
is sometimes diPcult to
distinguish ,et/een Arst and
second orders. # rate7
concentration graph li;e
those sho/n in Agure 1 /ill
sho/ the order /ith respect to
the reactant much more
clearly.
In rate la/sB ; is the rate constant of the reaction. %he value of k can ,e
considered constant for that reaction under those conditions. %he rate constant
is constant for a given reaction only if the concentration of reactants is ,eing
changed. 0o/everB k can vary ,ased on conditions such as temperatureB ionic
strength and the presence of catalysts. 1
; can ,e /or;ed out ,y rearranging the rate e4uation /hen the rate orders /ith
respect to ,oth catalase and hydrogen pero5ide are ;no/n>
"age = 1!
Figure 1(
%he concentrations of # and : have to
,e raised to some po/er to sho/ ho/
they aEect the rate of the reaction.
%hese po/ers are called the orders of
Figure 1-
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B ¿b
A ¿a¿
¿
k =rate¿
+igure 12 sho/s ho/ straight line plots can ,e used to determine the rate
constant depending on the order.
Figure 1/
%he general rate la/ for hydrogen pero5ide decomposition and catalase is k[H2O2 ] [catalase]. %he reaction is Arst order /ith respect to 0? and catalase as/ell. %he overall order of a reaction is given ,y (a @ ,) so the reaction is secondorder overall. %o sho/ this I /ill need to change the concentration of hydrogen pero5ide (/hilst;eeping the concentration of catalase constant) and measure ,y ho/ much theinitial rate increases or decreases. I can then determine the order /ith respect tohydrogen pero5ide. %he same can ,e done for varying concentrations of catalaseto And the order and hence k .
"age = 1$
Qeroth +irst ?rder
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#ims+rom my research I have found ho/ rate constants and orders can ,edetermined. :y carrying out the e5periment /ith an appropriate method andAnding the rate at various concentrations to And order and then k for eachtemperature. %his /ill allo/ me to use the #rrhenius e4uation /hich is
rearranged into a linear form /here I can calculate the activation energy as the
gradient is (− Ea
R ) /here R is the gas constant.
%o And <ma5B the velocity at diEerent hydrogen pero5ide concentration should ,efound. I /ill ,e a,le to plot initial velocity against su,strate concentration /hichshould give me the characteristic hyper,olic -ichaelis7-enten curve.
.ethods of ma7ing solutions
"ydrogen !eroxide solution
I /ill need to ma;e accurate solutions of various concentrations of hydrogen
pero5ide. %he concentration of hydrogen pero5ide is given as the volume of
o5ygen it produces9 1cm 3 of T166 volJ should produce 166cm 3 of o5ygen at S%".
# 6 vol concentration of hydrogen pero5ide is e4ual to 1. mol dm73. 1
<arious concentrations can ,e prepared as sho/n on the follo/ing ta,le>
Concentrati
on (vol)
<olume of 6 vol
hydrogen
"ero5ide (cm3)
<olume of distilled /ater
(cm3)
6 $6.6 6.6
1$ 12.$ *.$
16 1$.6 1$.6
$ *.$ 12.$
! $6.6 66.6
$.6 $.61 1.$ 3.$
"age = 1*
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&4uipment
• 6 vol hydrogen pero5ide
• Distilled /ater
• "ipette Aller
• $6cm3
volumetric Mas; and stopper• 16cm3 volumetric pipette
• $cm3 volumetric pipette
• 16cm3 measuring (mohr) pipette
• "asteur pipette
• +unnel
• 9a,els
-ethod
1. -easure re4uired volume of 6 vol hydrogen pero5ide /ith volumetric or
measuring pipettes. sing funnel ma;e sure all of it is transferred into $6cm3 volumetric Mas;
,y /ashing out /ith distilled /ater if needed3. #dd appropriate volume of distilled /ater to volumetric Mas; so meniscus
is aligned. se a pasteur pipette for the last fe/ drops.!. Replace stopper and turn ,ottle upside do/n so the solution is thoroughly
mi5ed$. 9a,el /ith appropriate concentration of hydrogen pero5ide
Catalase solution
Concentration (F) <olume of 166F catalase
solution (cm3)
<olume of distilled /ater
(cm3)166 166 626 26 6*6 *6 !6!6 !6 *66 6 26&4uipment
• 166cm3 volumetric Mas;
• "ipette Aller
• $6cm3 ,ea;er
• $cm3 volumetric pipette
• 16cm3 volumetric pipette
• "asteur pipette
• Stirring rod
• Solid catalase granules
• Distilled /ater
• Scales
• +unnel• 9a,els
"age = 1
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-ethod
1. "lace ,ea;er onto scaleB 'ero the reading and measure out 166g of
catalase. "our some /ater in ,ea;er and dissolve as much as possi,le /ith
stirring rod3. %ransfer all of this into a volumetric Mas;B /ash ,ea;er /ith more /ater
to get all into Mas;!. +ill /ith /ater so meniscus is on cali,ration line. se a pasteur pipette
for the last fe/ drops$. Replace stopper and turn ,ottle upside do/n several times to mi5
solution
%his is the 166F solution so to ma;e the others I /ill need to>
*. -easure out /ith pipette chosen volume of 166F solution into
volumetric Mas;. #dd distilled /ater until line is on meniscus2. Replace stopper and invert several times. 9a,el /ith appropriate concentration of catalase
!otassium +I:, .anganate
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.ethods of measuring rate %he e4uation for decomposition of hydrogen pero5ide /ith catalase is>
2 H 2
2
!2 H 2
+2 equation 1
%he rate of reaction is usually sho/n as the change in a property of reactant or
product per unit of time.
rate=change∈ prod"ct /reactant
time equation
%here are several /ays to measure the rate of reaction>
"roducts
• the volumes of gases (?) produced
• changes in pressure (o5ygen produced in a closed system creates
increasing pressure as reaction happens)Reactants
"age = 1
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• temperature change (as this decomposition is e5othermic)
• change in mass (as one of the product is o5ygen gas)
• p0 (p0 /ould drop if one of the reactants is an acid)
• concentration of 06 (this /ould drop as it converted into /ater and
o5ygen /hile reaction progresses)
-ethods I can use for my investigation are>
Gas syringe
%his is perhaps the easiest /ay to measure gas production. # gas syringe /ill
measure the volume of o5ygen produced from the reaction after catalase is
added.
&4uipment
• Clamp stand
• Hater ,ath
• 166cm3 gas syringe
• $6cm3 :Uchner Mas;
• 16cm3 measuring (-ohr) pipette
• Ru,,er ,ung
• "ipette Aller
• Ru,,er tu,ing
• Stop/atch
• 0ydrogen pero5ide solution
• Catalase Solution
-ethod
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Stop/atch
1. Clamp gas syringe hori'ontally to stand. -easure /ith measuring pipette $cm3 of hydrogen pero5ide solution into
:Uchner Mas;3. Connect tu,e to side arm and the other end to gas syringe!. Carefully place Mas; into the /ater ,ath and leave for 1 minute
$. "our catalase solution using measuring pipette into Mas; and 4uic;ly place,ung onto Mas;
*. S/irl mi5ture and start stop cloc;. Record the volume of gas at 16 second intervals for at least 3 minutes or
until reaction stops2. Clean conical Mas; and repeat 7* three more times /ith same
concentrations of hydrogen pero5ide and catalase. Repeat three times ,ut /ith diEerent hydrogen pero5ide and catalase
concentrations
Diagram>
Suba seal gas syringe
&4uipment
• Clamp stand
• Gas syringe
• 16cm3 syringe
• 0ypodermic needle
• $6cm3 :Uchner Mas;• Su,a seal
• Stop/atch
• Hater ,ath
• $cm3 volumetric pipette
• "ipette Aller
• 0ydrogen pero5ide solution
• Catalase solution
-ethod
1. Clamp gas syringe hori'ontally. Connect ru,,er tu,ing to side arm
"age = 1
:Uchner
Mas;
closed
/ith
,ung
containi
n 0?
Clam
Ru,,er
Hate
r
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Stop/atch
3. se volumetric pipette to put $cm3 of hydrogen pero5ide in the Mas;!. "lace su,a seal on :Uchner Mas;$. Carefully place Mas; in /ater ,ath and leave for 1 minute*. +ill syringe /ith catalase solution. "lace syringe through su,a seal and push do/n so all of the catalase is
transferred to Mas; ma;ing sure it is thoroughly mi5ed ,y s/irling2. Start stop cloc; and record the volume of gas in gas syringe at 16
second intervals for at least 3 minutes or until reaction stops. Hash outB dry Mas; and repeat 372 t/o more times /ith same
concentrations of hydrogen pero5ide and catalase16.Repeat three times ,ut /ith diEerent hydrogen pero5ide and catalase
concentrations
Diagram>
In5erted burette
%his /ill measure the volume of ? produced from the reaction in an inverted
,urette /hich /ill displace the /ater inside of it.
&4uipment
• Clamp stand
• :urette
• :o/lKplastic tu,
• Hater ,ath
• 16cm3 measuring pipette
• $cm3 volumetric pipette
• "ipette Aller
• $6cm3 :Uchner Mas;
• Ru,,er ,ung
• Ru,,er tu,ing
• Stop/atch
• 0ydrogen "ero5ide solutions
• Catalase solutions
-ethod
1. +ill ,urette /ith /ater and invert it into tu, of /ater.
"age =
16cm3
syringeAlled /ith
Clam
GasRu,,er
:Uchner
Mas;
containi
ng 0?
closed
/ith
Hate
r
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Stop/atch
. Secure /ith clamp and ma;e sure there is space for /ater to displace.
Release tap to remove any air ,u,,les and lo/er /ater /here initial
measurement can ,e made.3. -easure out hydrogen pero5ide solution (,ased on follo/ing ta,le as
,urette can only hold $6cm3) in volumetricKmeasuring pipette and pour in
:Uchner Mas;
Concentration (vol) -a5imum volume (ml)6 1$ 316 $$ 16! 1 $1 $6
!. #ttach tu,e to side arm and put the other end under /ater in ,uretteB
ma;ing sure it is secure$. Carefully place Mas; into /ater ,ath*. se measuring pipette to transfer catalase solution into :Uchner Mas; and
4uic;ly replace ,ung /hile s/irling. Start stop cloc; /hen Arst ,u,,le appears16.Record the volume at 16 second intervals for at least 3 minutes or until
reaction stops11.Hash and dry conical Mas;B reAll ,urette and repeat 17 t/o more times for
same concentrations1.Repeat three times ,ut /ith diEerent hydrogen pero5ide and catalase
concentrations
Diagram>
"age = 3
Inverted
,urette
Alled /ith
%ap
is:Uchner
Mas;
closed
/ith
,ung
containi
Clam
Ru,,er
Hate
r
%u, Alled
/ith
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TitrationI can And the e5act concentration of hydrogen pero5ide ,y a redo5 titration /ith
potassium manganate (<II) in presence of an acid. 3
-n?!7 @ $0 ? @ *0 @ L -n @ @ $? @ 20 ? equation $
:y adding an inhi,itor at certain times to stop the catalase from catalysing
hydrogen pero5ideB I can titrate that sample to determine the concentration of
0? at diEerent times and therefore the rate.
&4uipment
• Clamp stand
• ,urettes
• Hater ,ath
• +unnel
• $cm3 volumetric pipette
• 16cm3 volumetric pipette
• 16cm3 measuring pipette
• Conical Mas;
• 3 pipette Allers
•
$66cm3
,ea;er• * 166cm3 ,ea;ers
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• 0ydrogen pero5ide solutions
• Catalase solutions
• "otassium permanganate solution 6.6-
• Copper(II) sulphate solution
• &thanol
• Concentrated sulfuric acid• Stop cloc;
• Hhite tile
• 9a,els
-ethod for determining most eEective inhi,itor using gas syringe>
1. -easure out 6cm3 of hydrogen pero5ide into conical Mas;. #dd catalase and 4uic;ly replace ,ung3. Start timer and record volume of gas produced!. #t 16 seconds add the copper sulfate
$. Continue to record the volume for at least minutes*. Clean conical Mas; and dry. Repeat 17* for ethanol and sulphuric acid
%he ,est inhi,itor /ill 4uic;ly stop the reaction resulting in a smaller amount
of o5ygen produced.
-ethod>
1. -easure 1$cm3 of copper sulphate into 166cm 3 ,ea;ers /ith measuring
pipette. 9a,el ,ea;ers /ith times 167*6 seconds3. -easure out *6cm3 of hydrogen pero5ide solution into ,ea;er using
,urette!. #dd cm3 of catalase solution to ,ea;er /ith measuring pipette and start
stop cloc;$. &very 16 seconds using pipette transfer 16cm3 of reacting solution into a
,ea;er of the chosen inhi,itor (time intervals can vary depending on ho/
fast reaction seems to ,e ta;ing place)
No/ I have samplesB I need to titrate them /ith permanganate to And the
concentration of hydrogen pero5ide at each time
*. Rinse ,urette /ith potassium manganate (I<) allo/ing it drain into a /aste
,ea;er. Clamp ,urette to stand and All /ith the permanganate2. se volumetric pipette /ith pump to transfer ;no/n volume of
V4uenchedW solution to conical Mas;. #dd permanganate /hile s/irling the Mas;16.Slo/ do/n /hen pin; colour starts to appear11.Stop /hen pale pin; colour persists for 36 seconds1.Record the end point (volume of permanganate used)
13.Hash conical Mas; and repeat for other times1!.Repeat 17 1 t/o more times
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Stop/atch
1$.Repeat for diEerent concentrations of hydrogen pero5ide and catalase
Diagram>
"age = *
16cm3
volumetric
:ea;er
containin
ghydrogen
pero5ide
Inhi,itor
Conical Mas; /ith
sample of
V4uenchedW Hhite
:urette
/ith
Clam
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8is7 #ssessment
"age =
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"age = 2
Chemi
cal
"a*ard
symbol
8is7s !rotectio
n
!recautions ;5e
rall
8is7
0ydrog
en
pero5id
e
(6
volume)
1.1 Irritant
• irritatin
g to
eyes
and
s;in
• eye
protec
tion• gloves
• #void
contaminat
ion as this
may speed
up the
formationof o5ygen
and
pressure
,uild up
-edi
um
0ydrog
en
pero5id
e
(1$volume
and
less)
9o/
Catalas
e
(po/de
r)
1.
0armful
• harmful
as
po/derB
irritatin
g to
eyes• Irritatin
g to• eyes
and
s;in.• Ris; of
serious• damage
to eyes
• eye
protect
ion• use of
fume
cup,oard
• %o avoid
po/dered
en'ymes
escaping
into the air
s/itch oEthe fume
cup,oard
/hen carry
out the
/eighing so
particles
arenJt
inhaled
-edi
um
Catalas
e
(all
solutio
ns
a,ove
1F) Irritant
• irritant
for
solution
s
greater
than
1FB
irritatin
g toeyes
and
s;in• &n'yme
s are
sensiti'
• eye
protect
ion• gloves
• #void
contact
/ith s;in
and eyes
-edi
um
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General "recautions
• #void contact /ith s;in and eyes
• #void inhalation of vapour or mist
• Hear la, coat to avoid contamination of clothes
• Spillages should ,e ta;en care of immediately especially on Moor to avoidslips
• -ove seats under ,ench to prevent tripping around /or;ing area
• :e careful moving around and handling to prevent ,rea;ing glass/are
&mergency #ctions *
• Contact /ith s;in> /ash /ith soap and plenty of /ater
• Contact /ith eyes> Mood eyes /ith tap /ater for at least 1$ minutes
• S/allo/ed> rinse mouthB do not encourage vomiting• Inhaled> move into fresh air
"age =
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6ibliography
Introduction
1 0ydrogen "ero5ide general
http>KK,ro/nperiodf5hydrogenpero5ide.,logspot.co.u;K611K6Kthis7is7
/hat7hydrogen7pero5ide7molecule.html
Hhy is hydrogen pero5ide stored in dar; ,ottlesXhttp>KKhumantouchofchemistry.comK/hy7is7hydrogen7pero5ide7stored7in7
dar;7,ottles.htm3 %o5icology data net/or;B human health eEects> hydrogen pero5ide
http>KKto5net.nlm.nih.govKcgi7,inKsisKsearchKaX
d,s@hsd,>Yterm@YD?CN?@$!
! 0ydrogen "ero5ide 7 %o5icological ?vervie/
http>KK///.hpa.org.u;K/e,cK0"#/e,+ileK0"#/e,ZCK1!**6631$6
$ (Summary) !e "ecom#os$t$on o% Hy&ro'en erox$&e y *$+er atalaseB
[ohn HilliamsB -uspratt 9a,oratory of "hysical and &lectrochemistryB
niversity of 9iverpoolB 1 [ournal of General "hysiology
http>KK///.nc,i.nlm.nih.govKpmcKarticlesK"-C1!621KpdfK36.pdf
* Collision %heory
http>KKchem/i;i.ucdavis.eduK"hysicalZChemistryK8ineticsKRateZ9a/sKGasZ
"haseZ8ineticsKCollisionZ%heoryKCollisionZ%heoryZI %he &Eect ?f # Catalyst 7 Chapter 1!B section ! Chemistry ,y
:oundless
http>KK,oo;s.google.co.u;K,oo;sX
idfa3p####\:#[]printsecfrontcover]d4chemistry@,y@,oundless]
hlen]sa^]ei_2+*sjD?-ii6\H6ooGg#/]ved6CD&\*#&/##`v
onepage]4chemistryF6,yF6,oundless]ffalse
2 0omogeneous Catalysis 7 Chapter 13B section $ Chemistry ,y:oundless
http>KK,oo;s.google.co.u;K,oo;sX
idfa3p####\:#[]printsecfrontcover]d4chemistry@,y@,oundless]
hlen]sa^]ei_2+*sjD?-ii6\H6ooGg#/]ved6CD&\*#&/##`v
onepage]4chemistryF6,yF6,oundless]ffalse
%/o stage reaction proAle for a catalytic cycle
http>KK///.doc,ro/n.infoKpage63K#S#rates.htm
16 Some $oc!em$cal #ro#ert$es o% atalase %rom Ko!lra$B 0. %ayeA7
Nasra,adiB Department ?f :iochemistryB niversity of %a,ri'B 662
[ournal of :iological Scienceshttp>KK12.16.16!.132Kj,sK6K317[:S7#NSI.pdf
"age = 36
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11 Catalase> "roduct Information
http>KKhimediala,s.comK%DK%C63.pdf 1 "roposed -echanism of Catalase
http>KK,iology.;enyon.eduK:-:KChimeKcatalaseKframesKcatt5.htm`"ropos
ed -echanism of Catalase
13 &n'yme Catalysishttps>KK///.,oundless.comKchemistryKchemical7
;ineticsKcatalysisKen'yme7catalysisK1! Inhi,ition of Catalase #ctivity in Hines
http>KKajevonline.orgKcontentK*KK.a,stract1$ -athematics of #rrhenius &4uation
http>KKchem/i;i.ucdavis.eduK"hysicalZChemistryK8ineticsKReactionZRatesK
%emperatureZDependenceZofZReactionZRatesK#rrheniusZ&4uation
1* http>KKen./i;i,oo;s.orgK/i;iKStructuralZ:iochemistryK&n'ymeK-ichaelisZa
ndZ-entenZ&4uation
1 http>KK///.chemguide.co.u;KphysicalK,asicratesKorders.html`top
4ist of Figures
1 Four di<erent models used to represent a "; molecule
http=>>'''9'indo'suni5erse9org>physical?science>chemistry>h
ydrogen?peroxide9html #nthra4uinone "rocess for "roduction of 0ydrogen "ero5ide
http>KK///.loo;chem.comKChempediaKChemical7%echnologyKInorganic7
Chemical7%echnologyK2*3.html3 %he ?rientation of Collisions
http>KK///.chemguide.co.u;KphysicalK,asicratesKintroduction.html! #ctivation &nergy
http>KK///.chem.fsu.eduKchemla,Kchm16!*courseKactivation.html$ Catalysts or ma;ing it happen7 heterogeneous
http>KK///.spaceMight.esa.intKimpressKte5tKeducationKCatalysisKinde5.ht
ml
* Catalysis theory and practice
http>KK///.doc,ro/n.infoKpage6Kappendi5trans6*.htm
&n'ymes 7 .*. Induced7At model
http>KK///.clic;!,iology.infoKc!,KKpro.*.htm
2 Catalase Structurehttp>KKen./i;ipedia.orgK/i;iK+ile>CatalaseZStructure.png
On t!e -ec!an$sm o% t!e "ecom#os$t$on o% Hy&ro'en erox$&e y
atalaseB D 8eilin &+ 0artreeB niversity of Cam,ridgeB 13
http>KKrsp,.royalsocietypu,lishing.orgKcontentK1!K23K3.full.pdf 16 +actors aEecting &n'yme action
http>KK///.,iologymad.comKresourcesK&n'ymesRevision.pdf 11
1 +actors that aEect rate of reaction 7 temperature
http>KK///./e,chem.netKnotesKho/ZfarK;ineticsKrateZfactors.htm13 %he #rrhenius &4uation
http>KK///.chem.fsu.eduKchemla,Kchm16!*courseKarrhenius.html1! %he -ichaelis7-enten model
http>KKdepts./ashington.eduK/mat;insK;ineticsKmichaelis7menten.html
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1$ 9ine/eaver7:ur; plot
http>KKen./i;ipedia.orgK/i;iK9ine/eaverF&F26F3:ur;Zplot1* ?rders of reactions and rate e4uations
http>KK///.chemguide.co.u;KphysicalK,asicratesKorders.html1 Rate 9a/s
http>KK///.personal.;ent.eduKcearleyKgen$6Krevie/K;inetics.htm12 +igure 1!.1*B "roperties of Reactions %hat ?,ey Qeroth7B +irst7B and
Second7?rder Rate 9a/s
http>KK61,oo;s.lard,uc;et.orgK,oo;sKprinciples7of7general7chemistry7
v1.6mKs1276!7using7graphs7to7determine7rate.html
4ist of &quations
1 Septem,er 1$ issue of Chemistry Revie/
http>KK///.yor;.ac.u;KchemistryKschoolsKchemrevKprojectsKpero5ideK
E+olut$on o% atalases %rom Bacter$a to Humans -arcel Qamoc;yB "aul G.
+urtmUllerB Christian ?,ingerB niversity of Natural Resources and #pplied 9ife
SciencesB <iennaB 662http>KK///.nc,i.nlm.nih.govKpmcKarticlesK"-C$12*K
3
! #rrhenius e4uation deAnition
http>KKchemistry.tutorvista.comKinorganic7chemistryKarrhenius7e4uation.html
$ V%/o "oint +ormW of the #rrhenius e4uation
http>KK///.chem.fsu.eduKchemla,Kchm16!*courseKactivation.html
* Structural :iochemistryK&n'ymeK-ichaelis and -enten &4uation
http>KKen./i;i,oo;s.orgK/i;iKStructuralZ:iochemistryK&n'ymeK-ichaelisZandZ-
entenZ&4uation <ma5 and 8m can ,e determined ,y dou,le7reciprocal plotsB :iochemistry $th
&ditionhttp>KK///.nc,i.nlm.nih.govK,oo;sKN:8$$K
.ethods
1
0ydrogen "ero5ide VvolW
http>KKen./i;ipedia.orgK/i;iK0ydrogenZpero5ide`Decomposition
Catalase Solution
http>KK///.sigmaaldrich.comKtechnical7
documentsKprotocolsK,iologyKen'ymatic7assay7of7catalase.html
3
%itration
http>KK///.titrations.infoKpermanganate7titration7hydrogen7pero5ide
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8is7 #ssessment
1 1.1 http>KK///.jmloveridge.comKcoshKhydrogen
F6pero5ideF6*.pdf
0a'card $6http>KK///.cleapss.org.u;KattachmentsKarticleK6KCo
mpleteSet761376276.pdfX
SecondaryKScienceK0a'cardsK1. 0a'card 33
http>KK///.cleapss.org.u;KattachmentsKarticleK6KCo
mpleteSet761376276.pdfX
SecondaryKScienceK0a'cardsK1.3 0a'card 21
http>KK///.cleapss.org.u;KattachmentsKarticleK6KCo
mpleteSet761376276.pdfXSecondaryKScienceK0a'cardsK
Copper Sulfate
http>KK///3.ic,.usp.,rKcorpoeditorialK#R\I<?SKresiduosZ4uimicosKn
ovosKC?""&RF6S9+#%&F6S?9%I?NF6(CupricF6sulfate
F6standard).pdf
3 &thanol
http>KK///.sigmaaldrich.comK-SDSK-SDSKDisplay-SDS"age.doX
countryG:]languageen]productNum,er62],rand+98#]"
age%oGo%oR9httpF3#F+F+///.sigmaaldrich.comF+catalog
F+productF+Mu;aF+62F3+langF3Den! http>KK///.sigmaaldrich.comK-SDSK-SDSKDisplay-SDS"age.doX
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countryG:]languageen]productNum,er!3$$2],randSI#9]"
age%oGo%oR9httpF3#F+F+///.sigmaaldrich.comF+catalog
F+productF+sialF+!3$$2F3+langF3Den
$ https>KK/i;i.,ath.ac.u;Kdo/nloadKattachmentsK*13!2K9#:Z!7
hypodermics.pdf
* &mergency actionshttp>KK///.chem.uM.eduKfacilitiesKsafety7dealing7/ith7an7
emergency.shtml0a'ard http>KKchemistry.a,out.comKodKhealthsafetyKigK9a,oratory7Safety7
SignsKinde5.61.htm