Cleapps Recipe Book

The
CLEAPSS
Recipe Book

Introduction to this Edition
Recipe Cards were first produced in 1991 but very quickly demand led to an increase in the scope and
range of the information included. Over the ensuing 20 years we have added more and more
information whilst retaining the card format which users found useful.
This 2011 edition sees a change from cards to A4 sheets. The A5 card has become too small to contain
all the information required for some topics, and we believe that there is no further scope for reducing
the size of the print.
We have extended the number of separate entries from 74 to 106 to allow for more information on
some topics and to make other information easier to locate. We have added colour and some
photographs, where these are useful. The loose leaf A4 format also allows plenty of room for you to
insert recipes and instructions of your own.
Risk Assessment
Where a risk assessment includes control measures these have been incorporated into the instructions,
as describe in our Guidance Leaflet G90, Making and Recording Risk Assessments in School Science.
Each recipe therefore includes model risk assessments but does not include factors that you have to
thinks about, such as technician and teacher experience, and prep room conditions.
As in the previous edition, there is no mention of general bench solutions. In the past such solutions
were often much more concentrated than required and posed unnecessary hazards and risks. The
principle we follow (as in COSHH) is that the concentration of any reagent should be the lowest at
which the procedure works satisfactorily to give the intended result. Sometimes, therefore, we suggest
solutions which may appear to be an odd concentration but this nevertheless is the most suitable. For
example, 0.4 M sodium hydroxide solution is IRRITANT, whereas 0.5 M is CORROSIVE.
Making solutions
The recipes also make use of the laboratory jug as a measuring tool. Although apparently not
particularly accurate, we routinely achieve concentrations of between 1.95 and 2.05 M when using a jug
to prepare 2M sulfuric(VI) acid from concentrated sulfuric(VI) acid during the Practical Techniques in
Chemistry course. Clearly, using a jug can produce solutions with concentrations that are sufficiently
accurate for many laboratory purposes.
You will find more detail about making solutions in section 7.6 of the Handbook. If you run into
difficulties not covered in either the Recipe Book or the Handbook, phone CLEAPSS on 01895 251496
but do try the index first.

Contents
Recipe sheet Number
Agar 1
Alcohol/water and
propanone/water solutions
2
Alginate beads 3
Aluminium solutions 4
Ammonia gas 5
Ammonia solution (ammonium
hydroxide)
6
Ammonium chloride 7
Ammonium vanadate(V) solution 8
Azo dyes 9
Barium solutions 10
Benedicts qualitative reagent 11
Benedicts quantitative reagent 12
Biochemical indicators and tests 13
Bismuth nitrate(V) solution 14
Biuret reagent 15
Brodies fluid 16
Bromine water 17
Buffer solutions 18
Calcium chloride and nitrate(V)
solutions
19
Calcium hydroxide solution 20
Carbon dioxide 21
Cerium(IV) solutions 22
Chemiluminesence reactions 23
Chlorine gas 24
Chlorine water 25
Chromatography solvents and
locating agents
26
Chromium(III) chloride and
chrome alum solutions
27
Citric acid 28
Clock reactions

29
Recipe sheet Number
Cobalt(II) chloride solution and
thermochromic liquid
30
Copper(II) solutions 31
Crude oil alternative 32
2,4-Dinitrophenylhydrazine
solution
33
3,5-Dinitrosalicylic acid 34
Drosophila food base 35
Electroplating solutions 36
Enzymes 37
Etching solutions 38
Ethanoic acid 39
Fehlings solutions 40
Fixatives used before preserving
biological specimens
41
Gases less commonly used in
schools
42
Hydrochloric acid 43
Hydrogen gas 44
Hydrogen peroxide 45
Indicators (acid-base) 46
Indicator (universal) 47
Indicators (carbon dioxide) 48
Indicators for redox, precipitation
and complexometric titrations
49
Iodine solution 50
Iron(II) solutions 51
Iron(III) solutions 52
Lead(II) nitrate(V) 53
Lithium chloride 54
Magnesium sulfate(VI) 55
Manganese(II) sulfate(VI) 56
Mercury solutions 57
Methanal solution 58
Methanoic acid 59

Recipe sheet Number
Nickel sulfate(VI) 60
Nitric(V) acid 61
Nylon rope experiment 62
Oscillating reactions 63
Oxygen gas 64
Phosphoric(V) acid 65
Plant mineral requirement
solutions
66
Potassium and sodium
phosphates
67
Potassium chloride 68
Potassium chromate(VI) 69
Potassium dichromate(VI) 70
Potassium hydroxide 71
Potassium iodide 72
Potassium manganate(VII) 73
Preservatives used after fixing
biological specimens
74
Ringers and other saline
solutions for physiological use
75
Sandells solution 76
Silver nitrate(V) 77
Slime 78
Soap and bubble solutions 79
Sodium carbonate 80
Sodium chlorate(I) solution 81
Sodium chloride 82
Sodium ethanoate 83
Recipe sheet Number
Sodium hydrogencarbonate 84
Sodium hydroxide 85
Sodium silicate, the crystal
(chemical) garden and silicate
gels
86
Sodium thiosulfate 87
Stains for bacterial activity 88
Stains for cell contents 89
Stains for electrophoresis 90
Stains for fungal material 91
Stains for metabolic activity 92
Stains for plant material 93
Standard solutions for titration 94
Strontium chloride 95
Sulfur dioxide 96
Sulfur dioxide solution 97
Sulfuric(VI) acid 98
Testing for gases 99
Testing for negative ions 100
Testing for positive ions 101
Testing for organic functional
groups
102
Tin(II) chloride 103
Water (sea and hard) 104
Winklers method for dissolved
oxygen
105
Zinc sulfate(VI) 106
CLEAPSS Recipe Book
CLEAPSS 2011 1
1 Agar
For microbiological activities using purchased media, follow the instructions on the bottle.
The recipes are grouped into agars for microbiology and agars for other activities. All agars for
microbiological work need to be sterilised before and after use.

General Hazards Agar inhaled as a fine powder may cause an allergic reaction or other respiratory
problems. The use of agar that could isolate human pathogens (eg, blood agar) should
be avoided.
Control
measures
Use a balance in a non-working fume cupboard, ie, not switched on, with the sash
down, to weigh out agar. Use heatproof gloves to protect from scalding when handling
freshly-sterilised molten agar.

Procedure for
preparing
technical agar
(also called
agar-agar)
Mix 1.5 g of agar with 10 ml of water into a paste. Slowly add more water with stirring
until the volume is 100 ml. Heat the mixture with stirring on a boiling water bath to
95 C in the required container. This preliminary heating can be omitted if the agar is
going to be sterilised immediately, unless it is necessary to decant the agar into
smaller containers. In acid media, the amount of agar should be increased from 1.5 to
2 g. If the solidified agar in any recipe is too sloppy or too firm, repeat the procedure
using slightly more or less agar.
The agar gel is not stable in strongly alkaline solutions.
Usual sterilising
conditions
(If required) Autoclave the container(s) with the made-up suspension(s) for 15 minutes
at 15 psi (121 C).
Agars for microbiology
China blue lactose
agar
Use 3.6 g of China blue lactose agar powder in 100 ml of distilled water. If this mix
proves too thin for rough handling by students, then thicken by adding 0.5 g of agar-
agar (just thickener, no nutrients).
Crystal violet
agar. A selective
(against Gram
positive) medium
for soil bacteria
In a fume cupboard which is not switched on, add 0.005 g of crystal violet (HARMFUL,
DANGEROUS FOR THE ENVIRONMENT) to 1 litre of liquid nutrient agar solution. The
resultant solution is low hazard.
Glucose nutrient
agar
Add 0.5% w/v of glucose to molten nutrient agar.
Malt agar for fungi Mix 2 g of malt extract with 2 g of agar with 10 ml of water into a paste. Slowly add
more water with stirring until the volume is 100 ml. Autoclave the suspension at
10 psi (115 C) for 10 minutes.
Mannitol yeast
extract agar for
growing root
nodule bacteria
Mix 10 g agar in 1 litre of water, and dissolve in a boiling water bath. Add 0.5 g
K
2
HPO
4
, 0.2 g MgSO
4
.7H
2
O, 0.2 g NaCl, 0.2 g CaCl
2
.6H
2
O,10 g mannitol, and 0.4 g
yeast extract. Dispense as required and sterilise by autoclaving before use.
Nitrogen-free
mineral salts agar
for growing
nitrogen-fixing
bacteria
Dissolve 0.05 g FeCl
3
.6H2O in 500 ml distilled water. Add 2 g K
2
HPO
4
, 0.25 g
MgSO
4
.7H2O and 10 g glucose. Check the pH and adjust, if necessary, to 8.3 using
0.1 M NaOH. Pour into a bottle containing 1 g CaCO
3
and 7.5 g agar powder. Mix
and autoclave at 121 C for 20 minutes. Before pouring plates, swirl to thoroughly
mix the CaCO
3
and agar.
Nutrient agar for
bacteria
Mix 2 g of Bovril, 0.5 g of sodium chloride and 1.5 g of agar with 10 ml of water into
a paste. Slowly add more water with stirring until the volume is 100 ml. Heat/sterilise
the suspension as in Usual sterilising conditions above.
Starch malt agar
for growth of
fungi and
digestion of
starch
Mix 3 g of light malt powder (from home-brewing shops), 0.5 g of peptone (to
promote growth) in 20 ml of water. Also make a paste containing 1 g of soluble
starch in 10 ml of hot water. Add these two solutions to 1.5 g of agar with stirring and
slowly add more water with stirring until the volume is 100 ml. Stir before decanting
into smaller containers (if required) and sterilising. Autoclave the suspension at
10 psi (115 C) for 10 minutes.

CLEAPSS Recipe Book
2 CLEAPSS 2011
Agars for other activities
These should be made up as required, and not stored for long periods to avoid any unwanted microbial
activity. Dispose of as soon as possible after the activity. Sterilise in an autoclave any which are suspected of
microbial contamination.
Agar for starch
synthesis
Mix 0.5 g of glucose-1-phosphate and 1.5 g of agar with 10 ml of water. Slowly add
more water with stirring until the volume is 100 ml. Boiling, not sterilising, should be
sufficient.
Electrolytic agar Add one gram of sodium sulfate(VI) or other electrolyte to the hot agar solution
before pouring.
Ferroxyl agar gel
for rusting
experiments
Add 1.4 g of agar, 2 g of sodium chloride, 0.1 g of potassium hexacyanoferrate(III)
and 1 ml of phenolphthalein solution to the 100 ml of water and warm, with stirring, to
95 C. Pour the solution into Petri dishes.
Indicator agar Add 1 ml of the chosen indicator solution to the agar solution before pouring.
Mayonnaise agar
for lipase activity
(1)
Dilute 4 g salad cream or mayonnaise with 5 ml water and add 1 ml 0.1 M sodium
hydroxide solution (IRRITANT). Add about 1 ml of this alkaline mixture to a solution of
bromocresol green dye (about 0.003 g in 100 ml water) until the mixture just turns
blue-green and stir to ensure even distribution. Boil the resultant mixture with 2 g
agar, cool to 50-60 C then pour thin layers in Petri dishes. The plates will need to be
incubated at 30 C for 24 hours before being examined for orange-yellow areas
produced by lipases.
Mayonnaise agar
for lipase activity
(2)
Alternatively, mayonnaise agar can be made up without the dye and, after
incubation, the plates can be flooded with 0.4 M copper(II) sulfate solution and left for
30 minutes before being examined. Clear areas in the blue-green matrix indicate
where lipases have broken down the fatty acids in the mayonnaise.
Milk agar for
protease activity
Stir together 2 g low-fat milk powder (Marvel is recommended as it contains very little
fat), 1 g agar and 100 ml water. Heat as for technical agar and pour into Petri dishes
in very thin layers. Proteases should produce clear patches by breaking down
proteins in the milk within 30 minutes or so.
Phenolphthalein
indicator agar
Use 2 g agar in 100 ml boiling distilled (or deionised) water in a beaker. Add 10 ml of
0.2 M sodium carbonate solution and 5 ml of phenolphthalein into the beaker and stir
well.
Carbon dioxide in the atmosphere causes the colour to fade on storage, so the agar
is better prepared shortly before it is required.
Starch agar for
amylase activity
Mix a paste containing 1 g of soluble starch in 10 ml of cold water. Add 1.5 g of agar,
stir well and slowly add more water with stirring until the volume is 100 ml. Heat as
for technical agar above.
CLEAPSS Recipe Book
CLEAPSS 2011 3
2 Alcohol/water and propanone/water solutions
Quoted flash points appear to vary slightly between sources.
To prepare molar solutions, liquids can be weighed in tared containers.
2 M propanone solution is used in a rate of reaction experiment called the iodination of propanone.

To prepare 100 ml of an x% (v/v) solution of ethanol in water
Add x ml of ethanol to a 100 ml measuring cylinder.
Add water up to the 100 ml mark.
Label the solution. If it is highly flammable, then it needs the hazard warning label but, if flammable, a
label is not needed. However, the hazard classification needs to be written on any risk assessment.

% ethanol 0 10 20 30 40 50 60 70 80 90 100
Flash point (C) - 49 36 29 26 24 22 21 20 17 13
Density (g cm
-3
) at 20 C 1.00 0.98 0.97 0.96 0.95 0.93 0.91 0.89 0.86 0.83 0.78
Hazard - FLAMMABLE HIGHLY FLAMMABLE

To prepare 1 litre of 1 M methanol solution
Weigh out 32 g of methanol in a tared container or measure out 41 ml of methanol in a measuring
cylinder.
Add this to a 1000 ml measuring cylinder or 1 litre measuring jug.
Add water up to the 1 litre mark.
Label the solution harmful.

To prepare 1 litre of 1 M ethanol solution
Weigh out 46 g of ethanol in a tared container or measure out 58 ml of ethanol in a measuring
cylinder.
The solution is low hazard.

To prepare 1 litre of 2 M propanone solution
Weigh out 116 g of propanone in a tared container or measure out 147 ml of propanone in a
measuring cylinder.

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4 CLEAPSS 2011
3 Alginate beads
In studies of enzymes or the physiology of yeast cells, a valuable technique is to immobilise the enzyme or
cells inside beads of sodium alginate. The beads containing the enzyme/cells can then be used as usual or
packed into a column (eg, a syringe barrel) and a suitable substrate passed over them. The products are
collected at the bottom of the column and the immobilised enzymes or cells can be used again.
When making up the alginate and enzyme solutions it is essential to use purified water; otherwise calcium
ions in the water will cause the alginate to set prematurely.
Alginate beads can usually be stored overnight, covered and refrigerated but are unlikely to keep longer than
their non-immobilised components.
Always trial practicals to confirm activity of organisms or enzymes.

Preparing immobilised enzymes/cells in alginate beads
Make up a solution of the enzyme to be studied (see Recipe sheet 37 for enzymes), or a suspension
of yeast cells using purified water.
Sprinkle 2 g of sodium alginate in 100 ml of warm, purified water and mix using a mechanical stirrer.
Allow the solution to cool. Initially the mixture will form glutinous lumps, but it becomes smooth over
time.
Dissolve 3 g of calcium chloride-6-water in 200 ml of purified water in a 250 ml beaker.
Mix 2 ml of the enzyme/suspension with 8 ml of the 2% sodium alginate solution. Variations on these
proportions may be used.
Draw this up into a 10 ml syringe.
Add the sodium alginate/enzyme (or cell) mixture one drop at a time to the calcium chloride solution
making sure the tip of the syringe is held above the solution in the beaker.
Allow the beads to harden for a few minutes before straining them out of the beaker.

Hardened alginate beads in calcium chloride solution
CLEAPSS Recipe Book
CLEAPSS 2011 5
4 Aluminium solutions
Hydrated aluminium salts such as the chloride (AlCl
6
.6H
2
O, M = 241.5 g mol
-1
), sulfate(VI) (Al
2
(SO
4
)
3
.16H
2
O,
M = 630 g mol
-1
) and nitrate(V) (Al(NO
3
)
3
.9H
2
O, M = 375 g mol
-1
) absorb water (ie, they are hygroscopic) and
become damp on storage. Do check these chemicals before use.
Aluminium potassium sulfate(VI), also known as potassium aluminium sulfate, alum and potash alum is
easily stored and suitable for all activities where aluminium ions are required for testing. However, it is not all
that soluble in water, although it does make large octagonal crystals. Aluminium solutions are acidic.

Aluminium potassium sulfate(VI)
Formula: AlK(SO
4
)
2
.12H
2
O Molar mass: 474.39 g mol
-1
Solubility: 11 g per 100 ml

General Hazards
See Hazcard 2B. Aluminium salts in water are acidic.
Never use anhydrous aluminium chloride to make solutions. It reacts violently with
water producing toxic fumes of hydrogen chloride.

Mass (g) of solid to be used
Concentration
required
Volume (ml) of solution required Hazard warning
label
100 250 1000
0.01 M Ten-fold dilution of the 0.1 M solution -
0.1 M 4.74 11.86 47.44 -
0.2 M 9.49 23.72 94.88 -
Saturated (20 C) 12 29 114 -

Aluminium chloride (hydrated)
Formula: AlCl
3
.6H
2
-1

General Hazards
See Hazcard 2A. Aluminium salts in water are acidic.

Concentration
required
label
100 250 1000
0.1 M 2.41 6.03 24.14 -
0.5 M 12.07 30.18 120.72 -
Saturated (20 C) 90 225 900 IRRITANT

Preparing solutions of aluminium salts
Wear eye protection.
Measure out the indicated quantity of the aluminium salt.
Add the solid to about two thirds of the final volume of water in a beaker or laboratory jug.
Stir to dissolve, warming if necessary.
Pour the solution from the beaker into an appropriate measuring cylinder or laboratory jug and add
water to the required level.
Pour into a labelled bottle.
CLEAPSS Recipe Book
6 CLEAPSS 2011
5 Ammonia gas
Ammonia is less dense than air and very soluble in water so it has to be collected by the downward
displacement of air (upward delivery).
Theoretically, 2.1 ml of fresh concentrated (880) ammonia solution produces 1 litre of gas, although this is
never achieved in practice, so always use an excess. Older stocks of concentrated ammonia will be less
concentrated.
If the fountain experiment is to be carried out, a round bottom flask is substituted for the gas jar in the
diagram below. Many standard text books use a calcium oxide drying tube. There is no real need for this.

General Hazards See Hazcards 5 & 6. Ammonia begins to bubble off from 880 ammonia at about
55 C so heat gently and use anti-bumping granules to allow the ammonia to boil
off gradually.

Preparing ammonia gas
Use a fume cupboard. Wear goggles.
Add 10 ml of concentrated ammonia to the boiling tube
(CORROSIVE).
Set up the equipment as shown on the right.
Warm the boiling tube gently for about 5 minutes.
Knowing when the gas jar is full is not easy. Placing
moist red litmus at the neck of the inverted flask is not a
good indicator that the flask is full of gas. It is a matter of
judgement and experience. It might be better to use a
fresh boiling tube of concentrated ammonia for each gas
jar or flask required.
When the collection is finished, place a cover slip or,
better still, a large bung into the opening of the gas jar.
Store the gas jar upside down because ammonia is
lighter than air.
The residual ammonia solution can be poured down the
sink in a fume cupboard with plenty of water.

Heat
gently
10 ml of concentrated
ammonia solution
Inverted gas jar
held by a clamp
Anti-bumping
granules
CLEAPSS Recipe Book
CLEAPSS 2011 7
6 Ammonia solution (ammonium hydroxide)
The concentration of 35% (w/v) ammonia solution, (also known as 880 ammonia), is 18.1 mol dm
-3
.
Educational suppliers commonly supply this concentration although other concentrated solutions, eg,
25% (w/v) are available.
If kept for long periods, the concentration of ammonia solutions decreases because of leakage of gas from
the container.
If the concentrated ammonia is several years old it would be wise to test the concentration of the solution
before diluting further.
It is sensible not to store diluted ammonia solutions for long periods. It is better to prepare diluted solutions
only when required.
Solutions less than 1 M should be made by further dilution of 1 M ammonia solution and are best made fresh
before use.
If you have other concentrations of concentrated ammonia, then contact CLEAPSS for more advice. Some of
the values below are different from previous Recipe Cards as more precise information is now available.

Formula: NH
3
Molar mass: 17.03 g mol
-1
Solubility: infinite

General Hazards
Concentrated ammonia; see Hazcards 5 & 6.
Take great care when opening bottles of concentrated ammonia on hot days.

Volume of 880 ammonia to be used
Concentration
required
label
500 1000 2500
0.1 M Ten-fold dilution of the 1 M solution -
0.5 M Two-fold dilution of 1 M solution -
1 M 28 55 138 -
2 M 55 111 276 -
5 M 138 276 691 IRRITANT

Preparing ammonia solutions
Use a fume cupboard if concentrated ammonia or solutions more concentrated than 5 M ammonia are
used. Wear goggles (a face shield is preferable when handling large volumes) and chemical resistant
gloves.
Measure the indicated quantity of ammonia solution in an appropriate measuring cylinder.
Add the liquid to about two thirds of the final volume of water in an appropriate beaker or laboratory
jug.
Pour into a labelled bottle and mix well. Add a hazard warning if appropriate.
CLEAPSS Recipe Book
8 CLEAPSS 2011
7 Ammonium chloride
Formula: NH
4
Cl Molar mass: 53.5 g mol
-1

General Hazards See Hazcard 9A.

Concentration
required
label
100 250 1000
0.5 M 2.68 6.69 26.75 -
1.0 M 5.34 13.38 53.50 -
Saturated (20 C) 40 100 400 -

Preparing ammonium chloride solution
Measure out the indicated quantity of ammonium chloride.
CLEAPSS Recipe Book
CLEAPSS 2011 9
8 Ammonium vanadate(V) solution
Also known as ammonium polytrioxovanadate(V) and ammonium metavanadate.
Ammonium vanadate solution is used to illustrate the various oxidising states exhibited by vanadium.
Ammonium vanadate is not very soluble in pure water and has to be dissolved in alkali first, before making
acidic for use.

Formula: NH
4
VO
3
Molar mass: 117 g mol
-1
Solubility: 0.52 g per 100 ml

General Hazards Ammonium vanadate(V) is TOXIC. See Hazcards 9B, 91 and 98A.

Preparing 100 ml of 0.1 M ammonium vanadate solution
Wear goggles.
Dissolve 1.17 g of ammonium vanadate(V) in 20 ml of 2 M sodium hydroxide in a beaker. (The odour
of ammonia may be detected but it will cause no harm.)
Transfer the solution to a 100 ml measuring cylinder.
Add 1 M sulfuric(VI) acid to bring the total volume to 100 ml.
The yellow solution is irritant because of the presence of sulfuric(VI) acid.

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10 CLEAPSS 2011
9 Azo dyes
It is a myth that all azo dyes are carcinogens. The dyes can be prepared in schools as long as sensible
laboratory procedures are observed. On the whole, the more soluble a dye and the presence of sulfonic acid
groups on the benzene ring, then the safer the dye (eg, Orange II and methyl orange).
See Guide G195 for preparation of azo dyes from ethyl 4-aminobenzoate.
The recipes below can be used safely in schools.

General Hazards Sulfanilic acid (4-aminobenzenesulfonic acid), see Hazcard 4B. Sodium nitrate(III)
(sodium nitrite); see Hazcard 93. Naphthalen-2-ol; see Hazcard 70.
N,N-dimethylphenylamine; see Hazcard 4B. Sodium hydroxide; see Hazcard 91.
Ethanoic acid; see Hazcard 38A. Sodium carbonate; see Hazcard 95A.
Methyl orange; see Hazcard 32.

Preparing azo dyes as described is a two-stage process.
Stage 1: Preparing the diazonium salt of sulfanilic acid
Solution A:
Add to a boiling tube, 0.6 g of sulfanilic acid,
0.2 g of anhydrous sodium carbonate and 10 ml
of purified water.
Warm the mixture to boiling and then cool under
the tap. Add 0.3 g of sodium nitrate(III) and
agitate the test tube until the salt dissolves.
Solution B:
In another boiling tube, cool down 8 ml of
1 M hydrochloric acid using an ice-water
bath.
Add solution A to solution B. The diazonium salt settles out. It is more stable then many others and
will keep for some hours.
Test for the presence of free nitrous acid with starch-iodide paper. If the paper does not turn blue,
add a little more sodium nitrate(III). Swirl the contents of the boiling tube and divide the contents into
two test tubes.

Stage 2 (a): Preparing Orange II (IRRITANT) and dying cotton with it
Wear goggles. Wear disposable nitrile gloves.
In a test tube, dissolve 0.25 g of naphthalen-2-ol* in 4 ml of 1 M sodium hydroxide solution by
warming.
Cool the solution in an ice-water bath and pour it into a Petri dish.
Using forceps, add white cotton wool or cotton cloth (eg, bandage material) to the Petri dish to soak up
the solution.
Add the contents of the test tube containing the diazonium salt onto the cotton. The dye will appear.
Remove the cotton with forceps from the Petri dish and rinse it under cold water to remove any solid
dye. This is a soluble dye so do not do this for too long. Allow the cotton to dry.

* Other phenols, benzene diols, cresols and naphthols can be used with the diazonium salt of sulfanilic acid to produce other dyes.

Stage 2 (b): Preparing methyl orange (TOXIC)
Wear goggles. Wear disposable nitrile gloves.
In a test tube, add 0.2 ml of N,N-dimethylphenylamine to 0.2 ml of glacial ethanoic acid and agitate the
mixture well.
Add the contents of the test tube containing the diazonium salt to the test tube containing
N,N-dimethylphenylamine. Leave for 5 minutes for the red dye to form.
Add 2 ml of 2 M sodium hydroxide solution, heat to boiling and allow the solution to cool.
The solid orange sodium salt of methyl orange should form.

CLEAPSS Recipe Book
CLEAPSS 2011 11
10 Barium solutions
Purified water should be used to avoid cloudiness caused by the precipitation of barium sulfate(VI).
Barium chloride is classified as TOXIC if swallowed and barium nitrate(V) as HARMFUL if swallowed.
All solutions of barium nitrate(V) are LOW HAZARD.
Barium chloride is more soluble in water than barium nitrate(V).

General Hazards Barium chloride; see Hazcard 10A. Barium nitrate(V); see Hazcard 11.

Barium chloride
Formula: BaCl
2
.2H
2
-1

Concentration
required
label
100 250 1000
0.01 M Ten-fold dilution of 0.1 M solution -
0.1 M 2.44 6.11 24.43 HARMFUL
0.5 M 12.21 30.53 122.13 HARMFUL
Saturated (20 C) 36 90 260 TOXIC

Barium nitrate
Formula: Ba(NO
3
)
2
-1

Concentration
required
label
100 250 1000
0.1 M 2.61 6.53 26.14 -
Saturated (20 C) 9 45 90 -

Preparing barium salts solutions
Wear eye protection. Wear disposable nitrile gloves when weighing and preparing the solution.
Measure out the indicated quantity of the solid barium salt.
CLEAPSS Recipe Book
12 CLEAPSS 2011
11 Benedicts qualitative reagent
Benedicts solution or DNSA (see Recipe sheet 34) should be used in place of Fehlings solution to test for
reducing sugars because it is less hazardous.
Glucose, lactose and maltose are reducing sugars and give a positive test. Sucrose is a non-reducing sugar
and does not give a positive result.
Benedicts solution is less satisfactory in testing for non-reducing sugars and aldehydes in organic chemistry
in which case Fehlings or Sandells solutions will be needed.
No hazard warning symbol is required on the bottle as the concentrations of each of the constituents are low.
This solution is not suitable for colorimetric work. See Recipe sheet 12 for Benedicts quantitative reagent or
Recipe sheet 34 for DNSA.
To differentiate between reducing sugars, enzyme tests are required.

General Hazards Sodium carbonate; see Hazcard 95A. Copper sulfate(VI); see Hazcard 27C.

Procedure to produce 1 litre of solution
Measure out 170 g of trisodium citrate-2-water and 100 g of anhydrous sodium carbonate (or 256 g of
hydrated sodium carbonate).
Add the solids to about 850 ml of purified water in a 1 litre beaker.
Add 17.4 g of copper(II) sulfate(VI)-5-water and stir to dissolve.
Pour the solution into an appropriate measuring cylinder and dilute to the final volume.
Filter if necessary.
Pour into a labelled bottle and mix well.

Procedure for carrying out the test
The material under test is mixed with about 1 ml of water in a test tube or vial, and about 3 ml of
Benedict's reagent is added.
Place the test tube in a boiling water bath for about 5 minutes.
The colour should progress from blue (with no glucose present) to green, yellow, orange, red, and
then brick red or brown as glucose concentration increases.
CLEAPSS Recipe Book
CLEAPSS 2011 13
12 Benedicts quantitative reagent
The procedure detects the loss of blue colour as the sugar reduces the copper(II) ions to copper(I).
It can be used either in volumetric or colorimetric methods.
The addition of thiocyanate forms a complex and prevents the precipitation of copper(I) oxide.
See also Recipe sheet 34 for DNSA which is an alternative test for reducing sugars.

General Hazards Sodium carbonate; see Hazcard 95A. Copper sulfate(VI); see Hazcard 27C.
Potassium thiocyanate; Hazcard 95C. Potassium hexacyanoferrate(II) see
Hazcard 79. The volumetric method uses hot liquids.

Preparing 1 litre of Benedicts quantitative solution (BQS)
Measure out 75 g of anhydrous (or 160 g of hydrated) sodium carbonate, 200 g of trisodium citrate-2-
water and 125 g of potassium thiocyanate. Add these solids to 700 ml of boiled distilled water in a 1
litre beaker. Stir to dissolve, reheating if necessary.
Measure out 18 g of copper(II) sulfate(VI), dissolve in 100 ml of purified water and add this, with
constant stirring, to the solution made in step 1.
Add 0.25 g of potassium hexacyanoferrate(II) and pour the solution into a suitable volumetric flask.
Dilute to the final volume with purified water and mix well. For the colorimetric procedure below, dilute
35 ml of this solution to 100 ml with water. (No hazard warning label is required at these
concentrations.)

Using the solution: volumetric procedure
Place a 0.3% solution of glucose in a burette.
Place 10 ml of BQS into a 100 ml conical flask, heat to boiling and add about 2 g of anhydrous sodium
carbonate.
Add the glucose solution in 0.5 ml amounts, boiling each time until the blue or green colour just
disappears. Record the total volume of glucose added. When nearing the endpoint it is advisable to
boil for 30 seconds to avoid overshooting.
Repeat the procedure to confirm your result.
To measure the concentration of glucose in an unknown solution, repeat the procedure with glucose
solutions of unknown concentration.
By comparing the volume obtained on titration of the known and unknown samples, the %
concentration of the unknown can be calculated.
% conc o unknown solution =
u.S x B
A

A = titration volume for known glucose solution
B = titration volume for unknown glucose solution
(Some methods suggest adding a few drops of 1% aqueous methylene blue solution to act as an
indicator. It should be added when enough glucose solution has been added to turn the original
solution a very pale blue colour.)

Using the solution: colorimetric procedure
Place a 1% solution of glucose in a burette.
Place water in another burette and prepare a series of glucose solutions of varying concentration from
0 to 1%.
In a series of labelled fresh test tubes, add 1 ml of each solution prepared in step 2, to 10 ml of BQS.
Place these in a boiling water bath for a few minutes.
Allow the solutions to cool and any precipitates to settle.
Use a red or yellow filter in the colorimeter. Obtain the absorbance of light through each solution.
Use these readings to construct a glucose concentration calibration curve.
Use the calibration curve to identify unknown concentrations of glucose solution.

CLEAPSS Recipe Book
14 CLEAPSS 2011
13 Biochemical indicators and tests
Amino acid, polypeptide and protein tests
See also Recipe Sheet 15 for the Biuret test.
Coles
modification
of Millons
reagent
Test for
soluble
proteins
Wear goggles and gloves. Dissolve 10 g of mercury(II)
sulfate(VI) in 100 ml of 2 M sulfuric(VI) acid (solution A).
Separately, dissolve 1 g of sodium nitrate(III) (nitrite) in
100 ml of water (solution B). Before use, mix two volumes
of solution A with one of solution B. A reddish-brown
colouration or precipitate indicates the presence of soluble
proteins.
See Hazcards
62, 93, 98A.
Solution A and
the combined
solution are
TOXIC.
Marquiss
reagent
Test for
alkaloids
Wear goggles and gloves. Use a fume cupboard. Add 2
drops of 40% methanal solution to 3 ml of concentrated
sulfuric(VI) acid just before use. This is a spot test and
various colours appear. See
http://en.wikipedia.org/wiki/Marquis_reagent.
See Hazcards
63 and 98.
Label the
solution
CORROSIVE and
TOXIC.
Ninhydrin For amine
groups
Wear eye protection. Dissolve 0.1 g of ninhydrin in 100 ml
of water. A purple colour, known as Ruhemann's purple, is
produced when ninhydrin reacts with primary and
secondary amines, indicating the presence of amino acids.
Can be used as a spray or a dip, but needs to be heated, in
an oven at 110 C or with a hairdryer, for the colours to
appear.
See Hazcard
66. No hazard
warning is
required on the
solution.
Sakaguchi
test
Test for
proteins
containing
arginine
Wear goggles. Dissolve 5 g of sodium hydroxide in 100 ml
of water (solution A). Dissolve 1 g of napthalen-1-ol in
100 ml of water (solution B). For the test, one drop of
sodium chlorate(I) solution (10-14% available chlorine, see
Recipe sheet 81) is required as well. Proteins containing
arginine appear an intense red colour.
See Hazcards
70, 89 & 91.
Carbohydrate tests
See also 3,5-dinitrosalacylic acid (DNSA), Benedicts solution, Sandells solution and Fehlings solution.
Molischs
solution
For all
carbo-
hydrates
Wear goggles. Dissolve 5 g of napthalen-1-ol in 100 ml of
ethanol.
The solution containing a possible carbohydrate is
combined with a small amount of Molisch's reagent in a test
tube. After mixing, a small amount of concentrated
sulfuric(VI) acid is slowly added down the sides of the
sloping test tube, without mixing, to form a lower layer. Look
for a purple ring at the interface of the two layers.
See Hazcards
40A, 70 & 98.
Label the
solution HIGHLY
FLAMMABLE &
HARMFUL.
Periodic
acid Schiff
(PAS)
reaction
For
poly-
saccharides
Wear goggles and chemical-resistant gloves. Dissolve 1 g
of iodic(VII) acid (periodic acid) in 100 ml of water. Used in
conjunction with Schiffs reagent. Changes from colourless
to purple.
Iodic(VII) acid is
CORROSIVE &
OXIDISING. No
hazard warning
is required on
the solution.

CLEAPSS Recipe Book
CLEAPSS 2011 15
Cellular respiration
Janus
green B
Wear eye protection and disposable nitrile gloves when making up the
solution. Dissolve 0.3 g of the dye in 100 ml of purified water. Dilute this
solution ten times with water before use. Colour changes are from blue to
salmon pink.
See Guidance leaflet PS88 for more details including a practical activity.
No hazard
label is
required on
the solution.
Methylene
blue
Wear eye protection, and gloves to avoid staining the skin.
Dissolve 1 g solid in 100 ml water and add 0.6 g sodium chloride. The blue
indicator turns colourless as the dye is reduced.
See Hazcards
32 and 40. No
hazard label is
required on
the solution.
TTC Dissolve 1 g of 2,3,5-triphenyl tetrazolium chloride (TTC) in 100 ml of water.
(A 0.5% solution is less expensive and gives just as good results but takes
longer. It works well with maize seedlings.) Turns red as the dye is reduced.
Low hazard.
Tasting and genetics studies
Phenylthio-
carbamide
This is also called PTC, phenylthiourea or PTU. Dissolve 0.05 g in 100 ml of
water. Filter paper is soaked in the solution and then hung up to dry before
cutting into strips. See Handbook 15.13 for details on tasting investigations.
The solid is
VERY TOXIC;
see Hazcard
35. The
solution is low
hazard.
Vitamin C
DCPIP
solution
Also called 2,6-dichlorophenol indophenol, and phenol-indodichlorophenol.
Dissolve 0.1 g of dye in 100 ml of water. The standard vitamin C solution
should also be 0.1% (w/v). Add the blue dye until the colour does not
disappear.
Both are low
hazard
chemicals.
CLEAPSS Recipe Book
16 CLEAPSS 2011
14 Bismuth nitrate(V) solution
The solutions need to be kept acidic to avoid the formation of insoluble basic salts.

Formula: Bi(NO
3
)
3
.5H
2
O Molar mass: 395 g mol
-1

General Hazards See Hazcards 67 & 73B.

Preparing 100 ml of 0.1 M of bismuth nitrate(V) solution
Wear goggles.
Dissolve 5.34 g of bismuth(III) nitrate(V)5-water in 70 ml of 1 M nitric(V) acid.
Make up to 100 ml with 1 M nitric(V) acid.
Label the solution CORROSIVE.

CLEAPSS Recipe Book
CLEAPSS 2011 17
15 Biuret reagent
Biuret reagent tests for proteins.
Previous methods and recipes for this solution have used more concentrated solutions than that described
below. This meant that Y7 and 8 pupils were precluded from doing this experiment according to the advice
on Hazcard 91 (sodium hydroxide). However, the procedure below uses 0.1 M sodium hydroxide and
0.01 M copper(II) sulfate(VI) solutions and teachers might find that further dilution is possible.
It may be necessary to make acidic samples alkaline before the test for proteins is carried out.
Coomassie blue can also be used to test for proteins. It is available in a test kit with instructions for use. Its
advantages are that it does not require heating and is very sensitive.
Qualitative Biuret Reagent
This does not keep so only prepare what is required.

General Hazards Sodium hydroxide (solid) and 2 M solution. See Hazcards 91. Copper sulphate, see
Hazcard 27C.

Preparing 1 litre of Qualitative Biuret reagent
Wear goggles.
Weigh out 0.75 g of copper(II) sulfate(VI)-5 -water.
Prepare 1 litre of 2 M potassium or sodium hydroxide solution.
Dissolve the copper(II) sulfate(VI) in the alkali and label the solution CORROSIVE.

Procedure and use of Biuret solution suitable for Y7 and 8
Wear suitable eye protection.
Prepare 0.01 M copper(II) sulfate(VI) solution (Recipe Sheet 31).
Prepare 0.1 M sodium hydroxide solution (Recipe Sheet 85).
Place the sample of a liquid to be tested in a test tube to a depth of 10 mm.
Add the same volume of 0.1 M sodium hydroxide solution and agitate the test tube.
Add a few drops of the 0.01 M copper(II) sulfate(VI) solution and agitate the test tube.
A purple or pink colouration indicates the presence of protein.

Quantitative Biuret Reagent
This does keep.

General Hazards Sodium hydroxide (solid) and a 2 M solution see Hazcard 91. Copper(II) sulfate(VI)
see Hazcard 27C.

Wear goggles.
Dissolve 1.5 g of copper(II) sulfate(VI)-5-water crystals and 6 g of potassium sodium tartrate-4-water in
500 g of purified water.
Add 375 ml of 2 M sodium hydroxide with stirring.
If a precipitate occurs, add 1 g of potassium iodide.
Pour this mixture into a 1000 ml volumetric flask and dilute to 1 litre. Mix well. Label this solution
CORROSIVE as it is a 0.75 M sodium hydroxide solution.

For quantitative analysis, a series of standards can be produced with solutions of varying % dilutions of a
protein such as albumen, using a colorimeter with a 540 nm (green) filter. Unknown proteins solutions can
then be compared against these standards.

CLEAPSS Recipe Book
18 CLEAPSS 2011
16 Brodies fluid
Also known as manometer or manometric fluid.
Water strongly adheres to glass so requires an emulsifier to avoid water droplets sticking to the walls of the
manometer and affecting the readings, and to reduce capillary action in small bore manometers.

General Hazards Sodium azide is VERY TOXIC and contact with acids liberates a very toxic gas. (See
Hazcard 95B). For eosin, see Hazcard 32. Once in solution, the solution is LOW
HAZARD. Sodium azide may be omitted but the fluid will not keep as well and there
will be mould growth.

A. Preparing 1 litre of Brodies fluid
Weigh out 46 g of sodium chloride, 10 g of sodium tauroglycocholate (bile salts), 0.2 g sodium azide
(optional) and 0.5 g of eosin.
Make up to 1 litre with distilled or deionised water.

B. Brodies fluid substitute for short term use
44 g of sodium bromide.
1 g of liquid detergent (eg washing-up liquid).
0.3 g of Evans blue.

C. Simple version for short-term use
Use water with a food dye such as cochineal and a few drops of detergent.
CLEAPSS Recipe Book
CLEAPSS 2011 19
17 Bromine water
The solubility of bromine in water is 4 g, (ie, 1.25 cm
3
) in 100 g of water at room temperature. This would be
a 4% (w/v) solution. And its concentration would be 0.25 M.
Solutions equal to or greater than 0.06 mol dm
-3
(ie, a 0.3% v/v solution) are TOXIC.
Solutions equal to or more concentrated than 0.006 mol dm
-3
(0.1% w/v or 0.03% v/v) but more dilute than
0.06 mol dm
-3
should be labelled HARMFUL.
Aqueous solutions of bromine should be prepared just before use. If stored for long periods, especially
though the summer, the solution becomes paler as bromine vapour is lost.
Do not make this solution for the first time without seeking practical advice from a more-experienced
colleague.

General Hazards Bromine is VERY TOXIC and CORROSIVE (see Hazcard 15A). Hazcard 15B deals with
bromine water. 0.025 M bromine water has a considerable vapour of bromine gas
above it. It should be dispensed from a fume cupboard. More-dilute solutions can
be used in a well-ventilated room but staff should discourage any direct inhalation
of the vapour.
Sodium chlorate(I) is CORROSIVE (see Hazcard 89), 2 M hydrochloric acid; see
Hazcard 47A, Potassium bromate(V) is TOXIC, (see Hazcard 80).

Preparing 1 litre of 0.02 M solution (HARMFUL) of bromine water
Method 1
Use a fume cupboard. Wear goggles or a face shield and chemical-resistant gloves.
Using a disposable plastic pipette, add 1 ml of bromine to 500 ml of water in a 1 litre beaker.
Add a stirrer bar and place on a magnetic stirrer to dissolve the bromine. This can take over
20 minutes.
Dilute to 1 litre with water.
Alternatively, an ampoule containing 1 ml of bromine can be crushed under 500 ml of water, stirred
using a magnetic stirrer until it dissolves and made up to 1 litre with water. The solution must be
decanted so that no broken glass is present.

Method 2
Use a fume cupboard. Wear goggles or a face shield. Consider wearing gloves.
Dissolve 4.76 g of potassium bromide in 76 ml of water, add 14 ml of 10% (available chlorine) sodium
chlorate(I) solution (CORROSIVE) and 10 ml of 2 M hydrochloric acid (IRRITANT). Dilute to 1 litre with
water.

Method 3
This reaction is slow and it is better to leave the solution 24 hours before it is used.
Use a fume cupboard. Wear eye protection.
Add 1.12 g of potassium bromate(V) (TOXIC), 12 g of potassium bromide and 14 ml of 2 mol dm
-3

hydrochloric acid (IRRITANT) into a 1 litre jug or measuring cylinder.
Add water to 1 litre.
CLEAPSS Recipe Book
20 CLEAPSS 2011
18 Buffer solutions
Buffer solutions retain their pH on addition of small amounts of acid, alkali or on dilution.
Unless stated otherwise, buffers are low hazard but eye protection should be worn during preparation.
The pH value of a buffer is slightly altered by temperature but this is not usually significant. The values used
in the Recipe Sheet are for 20 C.
A definition for pH suitable for schools is minus the logarithm (to the base 10, ie log
10
) of the hydrogen ion
concentration in an aqueous solution. For a more advanced explanation, consult
http://en.wikipedia.org/wiki/PH.
There is also a pOH scale and pH + pOH =14.
A pH meter should be calibrated with standard buffers before it is used. These may be prepared from tablets
or commercial solutions.
Use distilled or deionised water to make up buffers.
Single-component buffers are very quick to make up but the values may not always be convenient.
The majority of buffers involve two components mixed in certain proportions. For accurate work, it is wise to
check their pH with a calibrated pH meter.
A universal buffer mixture can be used to obtain an array of buffer solutions from pH 2 to 11. The stock
solution can, usefully, be stored for several months.
Buffers (especially those between pH 4 and pH 7) do not keep well. Moulds develop which affect pH
readings and block the junctions on pH probes.
Commercial buffer solutions contain a mould inhibitor which allows longer storage.
To save time buffers can be stored as pre-weighed dry components or as frozen solutions. Before use, make
sure frozen solutions are returned to room temperature, that all components are fully dissolved, and check
the pH.
Special biological buffers (eg, TRIS) are available that do not use phosphate(V) or ethanoate ions which
might interfere with some biochemical processes. They are named Good buffers after their developer,
Norman Good. Different enzymes are inhibited by different reagents, so check the protocol and choose the
correct buffer system.
Single component buffers
pH 1.7
buffer
Dissolve 1.27 g of potassium hydrogen-ethanoate-1-ethanedioic
acid-2-water (potassium tetroxalate) in 10 ml of hot water and make
up to 100 ml with cold water.
Low hazard
See Hazcard 36A.
pH 4 buffer Dissolve 1.01 g of potassium hydrogenphthalate in 10 ml of hot
water and make up to 100 ml with cold water. The solution does not
keep well because of mould growth.
Low hazard
See Hazcard 13B.
pH 7 buffer Dissolve 0.77 g of ammonium ethanoate in 100 ml of cold water.
The solution does not keep well because of mould growth.
Low hazard
See Hazcard 9B.
pH 9.2
buffer
Dissolve 0.38 g of sodium tetraborate-10-water in 100 ml of water. Low hazard
See Hazcard 14.
pH 12.6
buffer
Use saturated limewater solution. Low hazard but wear
eye protection.
See Hazcard 18.

Temperature effects on single component buffers
Temperature / C 5 10 15 20 25 30 40 50 60 70 80 90
pH 4 buffer 4.01 4 4 4 4.01 4.01 4.03 4.06 4.09 4.12 4.16 4.2
pH 9.2 buffer 9.39 9.33 9.27 9.23 9.18 9.14 9.07 9.02 8.97 8.93 8.89 8.85

CLEAPSS Recipe Book
CLEAPSS 2011 21
Buffers for pH probe calibration
pH 4 buffer Dissolve 1.012 g of potassium hydrogenphthalate in 10 ml of hot
water and make up to 100 ml with cold water. The solution does not
keep well because of mould growth.
See Hazcard 13B
pH7 buffer 0.339 g of citric acid and 4.306 disodium hydrogenphosphate(V)-
12-water (or 1.707 g of anhydrous salt) dissolved in water and
made up to 100 ml with water.
See Hazcard 9B.
pH10 buffer 0.477 g sodium tetraborate-10-water and 18.3 ml of 0.1 M sodium
hydroxide (from a burette) is made up to 100 ml with water.
See Hazcards 14
and 91.
NB Although masses are given to 3 decimal places, it would be acceptable to work to 2 decimal places.

Double component buffers
To prepare these buffers, make up the individual components, then mix the quantities given for the required
pH. For accurate work, confirm the values with a calibrated pH meter and probe.

Using a solid and a solution.
General Hazards As 0.1 M sodium hydroxide solution is IRRITANT, wear eye protection when
preparing the buffers. All the resulting buffers are low hazard. See Hazcards 13B,
36C and 91.

3
10.21 g potassium hydrogen phthalate and 223 ml of 0.1 M
hydrochloric acid

Dilute each mixture to 1
litre solution with distilled or
deionised water.
4
10.21 g potassium hydrogen phthalate
5
10.21 g potassium hydrogen phthalate and 226 ml of freshly-made
0.1 M sodium hydroxide solution
6
6.81 g potassium dihydrogen phosphate and 56 ml of freshly-made
7
6.81 g potassium dihydrogen phosphate and 291 ml of 0.1 M sodium
hydroxide solution
8
6.81 g potassium dihydrogen phosphate and 467 ml of freshly-made
9
4.77 g sodium tetraborate-10-water and 46 ml of 0.1 M hydrochloric
acid
10
4.77 g sodium tetraborate-10-water and 183 ml of freshly-made 0.1 M
sodium hydroxide solution
11
2.1 g sodium bicarbonate and 227 ml of freshly-made 0.1 M sodium
hydroxide solution

CLEAPSS Recipe Book
22 CLEAPSS 2011
Mixing two prepared solutions Consult the relevant Recipe Sheets for the preparation of the solutions.
General
Hazards
All these buffers are low hazard. See relevant Hazcards for more information.

The citric acid/disodium hydrogenphosphate(V) buffer system
A is 0.2 M disodium hydrogenphosphate(V) and B is 0.1 M citric acid.
pH A (ml) B (ml) pH A (ml) B (ml) pH A (ml) B (ml)
2.2 2.00 98.00 4.2 41.40 58.60 6.2 66.10 33.90
2.4 6.20 93.80 4.4 44.10 55.90 6.4 69.25 30.75
2.6 10.90 89.10 4.6 46.75 53.25 6.6 72.75 27.25
2.8 15.85 84.15 4.8 49.30 50.70 6.8 77.25 22.75
3.0 20.55 79.45 5.0 51.50 48.50 7.0 82.35 17.65
3.2 24.70 75.30 5.2 53.60 46.40 7.2 86.95 13.05
3.4 28.50 71.50 5.4 55.75 44.25 7.4 90.85 9.15
3.6 32.20 67.80 5.6 58.00 42.00 7.6 93.65 6.35
3.8 35.50 64.50 5.8 60.45 39.55 7.8 95.75 4.25
4.0 38.55 61.45 6.0 63.15 36.85 8.0 97.25 2.75

The ethanoic acid/sodium ethanoate buffer
system
The ammonia/ammonium chloride buffer
system
Add the stated volume of A (0.2 M sodium
ethanoate solution) to the stated volume B
(0.2 M ethanoic acid)
Add the stated volume of A (0.2 M ammonia) to
the stated volume of B (0.2 M ammonium
chloride solution)
pH A (ml) B (ml) pH A (ml) B (ml)
3.8 12.0 88.0 8.4 12.5 87.5
4.0 18.0 82.0 8.6 18.5 81.5
4.2 26.5 73.5 8.8 26.0 74.0
4.4 37.0 63.0 9.0 36.0 64.0
4.6 49.0 51.0 9.25 50.0 50.0
4.8 60.0 40.0 9.4 58.5 41.5
5.0 70.5 29.5 9.6 69.0 31.0
5.2 79.0 21.0 9.8 78.0 22.0
5.4 85.5 14.05 10.0 85.0 15.0

The sodium hydrogencarbonate/sodium
hydroxide) system buffer system
The phosphate(V) buffer system. (potassium
salts may also be used)
Add the stated volume of A (0.1 M sodium
hydroxide) to 50 ml of 0.05 M sodium
hydrogencarbonate and dilute to 100 ml
(Wear eye protection.)
Add the stated volume of A [0.2 M sodium
dihydrogenphosphate(V)] to the stated
volume of B [0.2 M disodium
hydrogenphosphate(V)]
pH A (ml) pH A (ml) B (ml)
9.6 5.00 6.0 87.7 12.3
9.8 6.20 6.5 68.5 31.5
10.0 10.70 7.0 39.0 61.0
10.2 13.80 7.5 16.0 84.0
10.4 16.50 8.0 5.3 94.7
10.6 19.10
10.8 21.20
11.0 22.70
CLEAPSS Recipe Book
CLEAPSS 2011 23
Universal Buffer solutions

The Universal Buffer: Recipe 1
To prepare the stock solution, add 9.9 g of disodium hydrogenphosphate-12-water, (or 4.6 g disodium
hydrogenphosphate-2-water) (see Hazcard 72), 7.0 g of citric acid (see Hazcard 9) and 3.5 g of boric
acid (see Hazcard 14) to 250 ml of 1 M sodium hydroxide solution (see Hazcard 91) and dilute to
1000 ml with distilled/deionised water. This solution keeps well. Label the solution IRRITANT.

To make any buffer between 3.5 and 10
Place 0.1 M hydrochloric acid in a burette.
Place 20 ml of the stock solution in a 250 ml beaker on a magnetic stirrer. Clamp a pH probe into the
solution.
Add the acid to the solution in the beaker with constant stirring until the required pH is obtained. If too
much acid is added, use a little more stock solution to increase the pH value.
Add distilled/deionised water to make the final solution 100 ml.

The Universal Buffer: Recipe 2
To 500 ml of water, add 2 ml of concentrated ethanoic acid (see Hazcard 38A), 3 ml of 85%
phosphoric(V) acid (see Hazcard 72), and 2.4 g of boric acid (see Hazcard 14), and make the solution
up to one litre with distilled/deionised water.
Prepare 0.2 M sodium hydroxide solution (see Hazcard 91), and label the solution IRRITANT.

To make any buffer between 2 and 11
Put 0.2 M sodium hydroxide solution in a burette.
Place 100 ml of the acid solution in a 400 ml beaker on a magnetic stirrer. Clamp a calibrated pH
probe into the solution.
Add 0.2 M sodium hydroxide to the acid solution until the required pH is obtained.
Buffer solutions: Biological
Two component buffers are often used for biological systems. However, if an enzyme is affected by one, or
more, of the component ions, eg, phosphate(V), ethanoate or ammonium ions, specialist buffers are
required.
The cheapest specialist biological buffer is tris(hydroxymethylamino)methane (IRRITANT) known as TRIS. The
solutions made up from the data below are low hazard.
TRIS buffers
pH
Volume of 0.1 M
TRIS solution
Volume of 0.1 M
hydrochloric acid
7.0 100 93.2
7.5 100 80.6
8.0 100 58.4
8.5 100 29.4
9.0 100 11.4

A full range of biological buffers can be found at
www.sigmaaldrich.com/life-science/core-bioreagebnts/bioogical-buffers.html.
For products and applications such as extraction of materials or electrophoresis, see protocols at
www.ncbe.reading.ac.uk.
CLEAPSS Recipe Book
24 CLEAPSS 2011
19 Calcium chloride and nitrate(V) solutions
Hydrated calcium chloride and nitrate(V) absorb water from the atmosphere. On occasions, the solid
completely dissolves to leave a clear solution.
Distilled or deionised water should be used to make solutions. In hard water areas, solutions may be cloudy
if tap water is used.
Do not use anhydrous calcium chloride to make solutions.

General Hazards See Hazcard 19A

Formula: CaCl
2
.6H
2
-1

Preparing 100 ml of 0.1 M of calcium chloride solution
Dissolve 2.19 g of calcium chloride-6-water in 70 ml of water.
Make up to 100 ml with water.

Formula: Ca(NO
3
)
2
.4H
2
-1

Preparing 100 ml of 0.1 M of calcium nitrate(V) solution
Dissolve 2.36 g of calcium nitrate-4-water in 70 ml of water.
CLEAPSS Recipe Book
CLEAPSS 2011 25
20 Calcium hydroxide solution (Limewater)
Saturated calcium hydroxide solution, commonly called lime water, is a 0.02 M aqueous solution of calcium
hydroxide with a pH of 12.4.
It does not keep for long periods in large bottles because it reacts with carbon dioxide from the atmosphere.
It would be wise to start afresh each year.
Lime water, in small bottles designed for class use, will quickly cease to work. Class sets need to be tested
before handing out to the class.
1 M hydrochloric acid can be used to clean bottles that previously contained limewater.

Formula: Ca(OH)
2
-1

General Hazards Calcium hydroxide solid; see Hazcard 18. Splashed droplets of limewater in the eye
have caused quite severe irritation, so the solution should also be labelled and treated
as an IRRITANT even though strictly its dilution is such that it is not formally classed as
hazardous.

Preparing make 2.5 L of lime water in a bottle
Place 5 g of calcium hydroxide in a 600 ml beaker and half-fill it with water.
Stir the suspension and pour it via a funnel into a 2.5 litre bottle.
Fill the bottle with water, stopper and shake it.
Leave the bottle overnight for the suspension to settle.
When required, decant the limewater solution, slowly without agitating the sediment, into smaller
bottles for use in lessons.
Add more water and/or calcium hydroxide suspension when the level becomes low.

Preparing a large continual supply
If a large stock is required, keep an excess of calcium hydroxide in an aspirator protected by a soda-
lime tube, as shown below, and top up with distilled water as necessary.
Use about 100 g calcium hydroxide for a 10 litre aspirator. It might take a week to fully settle.

Absorption tube
filled with soda lime
Calcium
hydroxide
Lime
water

CLEAPSS Recipe Book
26 CLEAPSS 2011
21 Carbon dioxide
Carbon dioxide is prepared by the action of dilute hydrochloric or nitric acid on calcium carbonate, usually as
marble chips. If powdered calcium carbonate is used, the rate of gas production may be too rapid to be
easily controlled.
The gas is collected over water or by downward delivery. (Downward delivery uses a tube into the bottom of
an upright gas jar. It is also known as upward displacement of air and relies on the fact that carbon dioxide is
more dense than air.) Although carbon dioxide is slightly soluble in water (at room temperature, the solubility
of carbon dioxide is about 6.4 cm
3
of carbon dioxide per 100 ml of water), the rate at which it dissolves is
slow. However, if necessary, the gas can be collected over warm water in which it is less soluble.
Collection by downward delivery is quicker but it is difficult to ascertain exactly when the gas jar is full.
However, downward delivery is necessary for burning magnesium in carbon dioxide, in order to avoid water
interfering with the reaction.
Soda water is a saturated solution of carbon dioxide in water (carbonic acid).

General Hazards See Hazcards 20 & 47A.

In the 250 ml Bchner flask, add several lumps of marble chips.
Add enough water to immerse the base of the thistle funnel tube.
Set up the apparatus as shown in the diagram.
The inverted measuring cylinder should be full with water.
Now add 5 ml of 5 M hydrochloric acid (IRRITANT) down the thistle funnel.
Once the measuring cylinder is full of gas, it indicates the apparatus has been completely purged.
Remove it and replace with an inverted gas jar full of warm water.
Keep collecting gas jars of gas. When full of gas, either slide a gas jar cover in place or put a large
bung into the gas jar.
If the rate of gas production slows down but more gas is needed, add further 5 ml portions of 5 M
hydrochloric acid.

Add 5 mol dm hydrochloric acid
-3
250 ml measuring cylinder
Warm water
CLEAPSS Recipe Book
CLEAPSS 2011 27
22 Cerium(IV) solutions
Cerium(IV) compounds have a variable water content, which makes preparing accurate concentration
solutions very difficult. This is why the mass given in the recipe is approximate.
Cerium(IV) solutions, which have an intense yellow colour, are used in redox titrations.
Cerium(IV) solutions are more stable in solution than potassium manganate(VII) solutions.
Cerium(IV) solutions need to be prepared in dilute sulfuric(VI) acid.
[Hydrochloric acid must be avoided as cerium(IV) ions oxidise the chloride ions slowly to chlorine.]
Cerium(IV) solutions should be standardised against sodium ethanedioate solution before use.

General Hazards The solids are irritating to the eyes, respiratory system and skin.

Preparing 1 litre of 0.1 M cerium(IV) ions

Measure out about 64 g of ammonium cerium(IV) sulfate(VI)-2-water or 40 g of cerium(IV) sulfate(VI)-
4-water.
Add the solid to about 500 ml of 1 M sulfuric(VI) acid in a beaker.
Pour the solution into a 1-litre volumetric flask and make it up to the mark with water.

CLEAPSS Recipe Book
28 CLEAPSS 2011
23 Chemiluminesence reactions
Luminol (3-aminobenzene-1,2-dicarboxylic hydrazide or 3-aminophthalhydrazide) in alkaline solution is
oxidised to the 4-aminobenzene-1,2-dicarboxylate ion with the evolution of oxygen.
Goggles should be worn when preparing the solutions if using solid sodium hydroxide but spectacles are
suitable for the demonstrations using the solutions.
Recipe D is ideal for a chemiluminescent fountain demonstration using ammonia (see picture below).

Four recipes illustrating chemiluminesence reactions

A Solution A: Dissolve 0.4 g of luminol and 4 g of potassium hydroxide or sodium
hydroxide in 1000 ml of water. Label the container IRRITANT.
Solution B: Dilute 50 ml of fresh (10-14% available chlorine) sodium chlorate(I)
solution to 1000 ml. Label the container IRRITANT.
In a darkened room, mix equal volumes of solutions A & B together. The addition
of a pellet of potassium hydroxide or sodium hydroxide may produce more light.
See Hazcards
4B, 89, 91.
Wear goggles
to prepare the
solution.
B Dissolve 0.2 g of luminol and 1 g of potassium hydroxide or sodium hydroxide in
1000 ml of water. To a known volume of this solution add an equal volume of
20 vol hydrogen peroxide solution.
In a darkened room, sprinkle a few crystals of potassium hexacyanoferrate(III)
onto the surface of the solution and observe the trails of light.
The addition of a pellet of potassium hydroxide or sodium hydroxide may
produce more light.
See Hazcards
4B, 50, 79, 91.
Wear goggles
to prepare the
solution.
C Solution A: Dissolve 0.2 g luminol, 4 g of anhydrous sodium carbonate, 24 g of
sodium hydrogencarbonate, 0.5 g of ammonium carbonate and 0.4 g of hydrated
copper(II) sulfate(VI) in 1000 ml water.
Solution B: Prepare 1000 ml of 5 vol hydrogen peroxide solution.
Mix equal volumes of A and B in a dark room or in a box with spy holes.
See Hazcards
4B, 27C, 50,
95A.
D Solution A: Mix together 0.2 g luminol, 11 g of anhydrous sodium carbonate,
8 g of sodium hydrogencarbonate, 0.5 g of ammonium carbonate and 0.4 g of
copper(II) sulfate(VI) in 1000 ml water. Add 25 ml of 10 vol hydrogen peroxide.
Solution B: Dissolve 0.1 g of cobalt(II) chloride and 0.1 g of sodium nitrate(III)
(ie, sodium nitrite) in 1000 ml of water.
Mix equal volumes of A and B in a dark room or in a box with spy holes.
See Hazcards
4B, 9A, 25,
27C, 50, 89, 93,
95A.

The chemiluminescent fountain demonstration
The flask is originally filled with ammonia. Two containers labelled A and B which contain 1 litre each of
solution A & B in Recipe D are used to supply the solution in the fountain. The solutions are connected via a
T-piece into a single tube which is inserted into the flask.
CLEAPSS Recipe Book
CLEAPSS 2011 29
24 Chlorine gas
Version 1: Concentrated hydrochloric acid on potassium manganate(VII)
The reaction of concentrated hydrochloric acid on potassium manganate(VII) is the more common and
reliable of the two methods. However it uses more-hazardous starting chemicals.
It produces chlorine quickly.
The glassware is difficult to clean at the end and more-concentrated hydrochloric acid is required to remove
manganese(IV) oxide stains.
colleague.

General Hazards See Hazcards 22A, 20 & 81. Double check that you are using concentrated
hydrochloric acid and not concentrated sulfuric(VI) acid. Explosions have resulted from
using the wrong acid.

Use a fume cupboard. Make sure it is working. Wear goggles.
Set up the equipment as in the illustration below. Make sure that the thistle funnel tube nearly reaches
the base of the flask.
Place at least 5 g of potassium manganate(VII) (OXIDISING & HARMFUL) in the Bchner flask.
Pour the concentrated hydrochloric acid (CORROSIVE) down the thistle funnel.
Collect the gas in a gas jar. When the contents of the gas jar are clearly green, assume the gas jar is
full. Or (optional), plug the gas jar with a large wad of mineral wool, or cotton wool, and put some
damp blue litmus paper on top. It takes time for the chlorine to diffuse through the wool and bleach the
litmus. When the bleaching is complete the jar will be full. Remove it and replace the mineral wool with
a gas jar cover or a large bung.
More gas jars of gas can be collected.
To dispose of the reaction mixture, pour it down the fume cupboard sink with lots of water. The stained
glass may need treating with a little concentrated hydrochloric acid before washing further.

Mineral wool
Moist blue litmus
At least 5 g of
Potassium
Manganate(VII)
Wooden block
Gas jar
Delivery tube as far
down as possible
In a fume cupboard,
pour concentrated
hydrochloric acid down
the thistle funnel
Bchner flask
CLEAPSS Recipe Book
30 CLEAPSS 2011
Version 2: 5 M hydrochloric acid added to sodium chlorate(I) solution
This reaction is slower than first version and is easier to control. The glassware is easy to clean at the end of
the procedure.
Sodium chlorate(I) solution is difficult to store and must be less than 1 year old to obtain a good supply of
chlorine. Domestic bleach is not suitable.
As the reaction subsides, either the acid or the sodium chlorate(I) solution can be alternatively added to the
reaction vessel to supply more gas.
The gas preparation arrangement (shown below) is known as the Andrews method. It is an alternative
arrangement to using a thistle funnel. The arrangement in version 1 can also be used.
colleague.

General Hazards See Hazcards 22A, 20 & 89. Sodium chlorate(I) is a solution provided by the
supplier often as sodium hypochlorite. Sodium chlorate(V) is a solid. Do not get
them mixed up.

Place at least 50 ml sodium chlorate(I) solution (CORROSIVE) in the Bchner flask.
Set up the equipment as in the illustration below. The magnetic stirrer is optional but it does appear to
make the procedure more efficient.
Pour the 5 M hydrochloric acid (IRRITANT) into the separating funnel.
Turn the stirrer on and drip the acid into the flask. The solution will now bubble as the chlorine is
produced.
Collect the gas in a gas jar. When the contents of the gas jar are clearly green, assume the gas jar is
full. Or (optional), plug the gas jar with a large wad of cotton or mineral wool and put some damp blue
litmus paper on top. It takes time for the chlorine to diffuse through the cotton wool and bleach the
litmus. When the bleaching is complete the jar will be full. Remove it and replace the mineral wool with
a gas jar cover or a large bung.
To dispose of the reaction mixture, pour it down the fume cupboard sink with lots of water.

Separating funnel
5 M hydrochloric acid
Magnetic stirrer
Sodium
chlorate(i)
solution
Mineral wool
Moist blue litmus
Gas jar
Delivery tube as far
down as possible
Rubber tubing
CLEAPSS Recipe Book
CLEAPSS 2011 31
25 Chlorine water
The solubility of chlorine in water is about 0.6 g in 100 ml of water at room temperature. This would be a
0.6% (w/v) solution. Its concentration would be 0.085 M.
An aqueous solution of chlorine should be prepared just before use. It does not keep for more than just a few
days and should not be stored.
It is very difficult to make up solutions of known concentration. All that is required for displacement reactions
is that the solution works. It should, therefore, be trialled before use.
Do not make this solution for the first time without seeking practical advice from a more experienced
colleague.

Three methods are described below.

General Hazards Chlorine gas is TOXIC and CORROSIVE (see Hazcard 22A). Hazcard 22B deals with
chlorine water. Freshly-made chlorine water is formally LOW HAZARD but it has a
considerable vapour of chlorine gas above it. It is better dispensed from a fume
cupboard. More dilute solutions can be used in a well-ventilated room but staff
should discourage direct inhalation of the vapour.

Method 1
Bubble chlorine gas into 250 ml water in a gas jar until the solution goes light green. Use the
equipment for preparing chlorine gas but fill the gas jar half-full with water.
Method 2
Sodium chlorate(I) solution is an aqueous solution. Do not become confused with sodium chlorate(V), which
is a solid.
10% w/v available chlorine sodium chlorate(I) solution does not store well. Over two years it may become
completely useless.
Place 10 ml of 10% w/v available chlorine sodium chlorate(I) solution (CORROSIVE) in 1 litre beaker.
Add about 80 ml of water and 10 ml of 2 M hydrochloric acid. Stir well.
Dilute to a suitable volume with more water.
Method 3
Sodium dichloroisocyanurate is used for purifying water.
Use a fume cupboard. Wear eye protection.
Add 3 g of sodium dichloroisocyanurate (OXIDISING; HARMFUL), to 100 cm
3
of water. When the solution
is clear, add 100 cm
3
of 1 mol dm
-3
hydrochloric acid.

CLEAPSS Recipe Book
32 CLEAPSS 2011
26 Chromatography solvents and locating agents
A mixture of compounds is placed on a stationary phase such as paper, or silica on thin layer plates.
Chromatography involves passing a solvent through the stationary phase which separates the components
of the mixture due to subtle differences in a compound's partition coefficient between the mobile solvent and
the stationary phase.
A locating agent is used to emphasise where the components of the mixture end up after the
chromatography has finished. This is particularly important for colourless components of mixtures.

Control measures
for all of the
following solutions
Solutions are better made up in a fume cupboard.
There must be no sources of ignition in the vicinity.

Substances
separated

Amino acids Solvent: Butan-1-ol, glacial ethanoic acid, water; 6:1:2 by
volume.
Locator: Apply ninhydrin and heat in an oven at 110 C or with a
hairdryer.
See Hazcards 38A,
66, 84B. Label
solvent HARMFUL and
CORROSIVE.
Analgesics,
eg, aspirin,
paracetamol
Solvent: Ethyl ethanoate, hexane, ethanoic acid; 10:9:1 by
volume.
Locator: UV light or iodine vapour. Do not look directly at UV
radiation sources.
Label the solvent
HIGHLY FLAMMABLE
and CORROSIVE. See
Hazcards 43A, 54A.
Anthocyanins
(plant
pigments)
Solvent: 100 ml 50% aqueous methanol solution with 1 ml of
ethanoic acid or
1 ml concentrated hydrochloric acid in 100 ml ethanol.
Locator: Natural colour. (The components are light sensitive so
run the chromatograms in the dark if possible and
quickly note or photograph the results.)
Label the solvent
HIGHLY FLAMMABLE
and TOXIC. See
Hazcards 38A, 40B.
Inks from
Biro pens
Solvent: Butan-1-ol, ethanol, water; 3:1:1 by volume. The
addition of a few drops of 880 ammonia is said to
produce a better chromatogram.
Locator: Natural colour.
Label the solvent
HIGHLY FLAMMABLE
and HARMFUL. See
Hazcards 6, 40A,
84B.
Chlorophyll Solvent: Propanone, petroleum spirit (100-120 C); 1 :9 by
volume or
cyclohexane, propanone, ethoxyethane; 2:1:1 by
volume.
Locator: Natural colour of dyes and UV light. Do not look directly
at UV light. (The components are light sensitive so the
results should be noted or photographed quickly.)
Label the solvents
HIGHLY FLAMMABLE.
See Hazcards 45A,
85 or 42, 45B, 85.
Lipstick Solvent: 3-Methylbutan-1-ol,propanone and water; 2:1:9 by
volume.
Locator: Natural colour.
The solvent is LOW
HAZARD See
Hazcards 84C, 85.
Metal ions Solvent: Propanone, hydrochloric acid (conc), Distilled
water; 17:2:1 by volume.
Locator: Use conc ammonia solution followed by 0.1% dithio-
oxamide (rubeanic acid).
Label solvent HIGHLY
FLAMMABLE and
IRRITANT Label the
locator CORROSIVE.
See Hazcards 35,
47A, 85.
Nitration of
methyl
benzoate
For separating the crude products obtained from the nitration of
methyl benzoate.
Solvent: Ethoxyethane, pet ether (80/100 C); 1:9 by volume.
Locator: UV light. Do not look directly at UV radiation sources.
Label the solvent
HIGHLY FLAMMABLE
and HARMFUL. See
Hazcards 42, 45A.
Sugars Solvent: Ethyl ethanoate, pyridine, water, 8:2:1 by volume.
Locator: Dab Benedicts solution on the chromatogram and dry in
an oven at 110 C.
Label the solvent
HIGHLY FLAMMABLE.
See Hazcards 4C, 8,
43A.
CLEAPSS Recipe Book
CLEAPSS 2011 33
27 Chromium(III) chloride and chrome alum solutions
The colour of chromium(III) solutions varies with temperature and storage.
Use cold water to dissolve the salt and do not heat the solutions.
Prepare just before use.

General Hazards Chromium (III) salts are IRRITANT. See Hazcard 24.

Formula: CrCl
3
.6H
2
-1

Preparing 100 ml of 0.1 M chromium(III) chloride solution
Dissolve 2.67 g of chromium(III) chloride-6-water in 70 ml of cold pure water.
Make up to 100 ml with pure water.

Formula: CrK(SO
4
)
2
.12H
2
-1

Preparing 100 ml of 0.1 M chromium(III) potassium sulfate(VI) solution
Dissolve 5.0 g of chromium(III) potassium sulfate(VI)-12-water in 70 ml of cold pure water.

CLEAPSS Recipe Book
34 CLEAPSS 2011
28 Citric acid
Also known as 2-hydroxypropane-1,2,3-tricarboxylic acid.
It can purchased in either an anhydrous or monohydrate form.
Values in italics below are for the monohydrate form.

General Hazards See Hazcard 36C.

Formula: C
6
H
8
O
7
-1
Formula: C
6
H
8
O
7
.H
2
-1

Concentration
required
label
100 250 1000
0.1 M 1.92 (2.10) 4.80 (5.35) 19.21 (21.01) -
0.5 M 9.60 (10.51) 24.02 (26.27) 96.06 (105.07) -
1.0 M 19.21 (21.01) 48.03 (53.54) 192.12 (210.14) -

Procedure
Measure out the indicated quantity of the solid citric acid.

CLEAPSS Recipe Book
CLEAPSS 2011 35
29 Clock reactions
These are used for rate of reaction investigations.
Solutions should be made a day before use and stored in the place where they will be used to attain room
temperature.
Clock reactions need to be tried out beforehand so that concentrations can be tweaked to give reasonable
times. Summer and winter temperature variations can also alter reaction times.
The suggested amounts are for student activities. These reactions, however, make excellent demonstrations
and the volumes can be significantly increased.
The asterisk by the name of a solution indicates that this reagent is involved in the main reaction and its
concentration can be varied. However, the amount of each solution must remain as specified.
Varying reaction temperature will alter the time lapse for the clock.
The addition of transition metal ions, eg, copper(II) ions, as a catalyst can also be investigated. However, you
need to trial the experiment as the sequence that components are added is very important.

The hydrogen
peroxide/potassium
iodide reaction
(The Harcourt-
Essen reaction)
Solution A 1% starch solution (indicator)
Solution B* 4 vol hydrogen peroxide
Solution C* 0.1 M hydrochloric acid
Solution D 0.01 M sodium thiosulfate
Solution E* 0.05 M potassium iodide
Add 5 drops of solution A to a suitable beaker plus 10 ml
each of solutions B to E in that order.
See Hazcards 47A
& B, 67, 95C.

Wear eye
protection.
The thiosulfate/acid
reaction
(disappearing
cross)
Solution A 0.1 M sodium thiosulfate
Solution B 1 M hydrochloric or 0.05 M sulfuric(VI) acid
Add 1 ml of the acid for every 10 ml of sodium thiosulfate.
Once the run is complete, dispose of the contents in 0.5 M
sodium carbonate solution to stop the reaction and neutralise
sulfur dioxide.
See Hazcards 47A,
95C & 98.

Wear eye
protection. Do not
use temperatures
above 50 C.
The potassium
iodate/sodium
metabisulfite
reaction
(The Landolt iodine
clock)
Solution B* 0.025 M potassium iodate(V) solution (5.35 g
per 1000 ml of solution)
Solution C* 0.025 M sodium metabisulfite solution (4.75 g
per 1000 ml of solution. Prepare in a fume cupboard.)
Add 5 drops of solution A to a suitable beaker plus 10 ml
each of solutions B and C.
See Hazcards 80,
92.

Wear eye
protection.
The reaction
between iron(III)
ions and iodide
ions
Solution B 0.01 M sodium thiosulfate
Solution C* 0.025 M iron(III) chloride using 0.1 M
hydrochloric acid to dilute a more concentrated iron(III)
solution
Solution D* 0.025 M potassium iodide
Mix 5 drops of solution A followed by 10 ml each of solutions
B, C and D in that order.
See Hazcards 41,
47B, 55, 67, 95C.

Wear eye
protection.
The potassium
iodide/potassium
persulfate reaction
Solution A Dissolve 0.25 g of sodium thiosulfate in 100 ml
of 1% starch solution
Solution B* Dissolve 13.50 g of potassium persulfate in
1000 ml of solution
Solution C* 0.1 M potassium iodide
Mix 5 ml of solution A with 50 ml each of solutions B and C in
that order.
See Hazcards 47B,
95B, 95C.

Wear eye
protection.

CLEAPSS Recipe Book
36 CLEAPSS 2011
30 Cobalt(II) chloride solution and thermochromic liquid
General Hazards Cobalt salts are TOXIC (See Hazcard 25). Reported carcinogenic and sensitisation
warnings about cobalt compounds have become more severe in recent years.
CLEAPSS has not received any reports from educational establishments of cobalt
salts causing harm. The concerns for health are in mining and metallurgical
applications used on a daily basis and in large doses.

Formula: CoCl
2
.6H
2
-1

Preparing 100 ml of 0.1 M cobalt(II) chloride solution
Wear eye protection. Wear disposable gloves and consider weighing the solid in non-working fume
cupboard with the sash window low enough to stop any particles being inhaled.
Dissolve 2.38 g of cobalt(II) chloride-6-water in 70 ml of water.
The solution should be labelled TOXIC.

Formula: Co(NO
3
)
2
.6H
2
-1

Preparing 100 ml of 0.1 M cobalt(II) nitrate solution
Use the method above but dissolve 2.91 g of cobalt(II) nitrate-6-water in 70 ml of water.

Formula: CoSO
4
.7H
2
-1

Preparing 100 ml of 0.1 M cobalt(II) sulfate solution
Use the method above but dissolve 2.81 g of cobalt(II) sulfate(VI)-7-water in 70 ml of water.

Thermochromic liquid
Used in models of hot-water systems.

Dissolve 40 g of hydrated cobalt(II) chloride in 1 litre of ethanol without heating.
Add drops of water and stir until pink. A drop or two of ammonia solution will precipitate the hydroxide
which shows the flow in the model.
Label the solution TOXIC & HIGHLY FLAMMABLE.
CLEAPSS Recipe Book
CLEAPSS 2011 37
31 Copper(II) solutions
It is advisable to use distilled or deionised water to make these solutions.
Solutions of copper(II) salts are sometimes cloudy. If this is the case, add 1 ml of 1 M sulfuric acid and stir.
Continue this procedure until the solution is clear.
The Health & Safety Executive provide hazard classifications for solutions by % (w/v). We generally measure
concentration in mols per dm
-3
rather than % (w/v), which leads to inconsistencies in hazard classification
between different copper salts with the same concentration. In the tables below the hazard classifications in
brackets are consistent with that for copper sulfate(VI), the most commonly used copper(II) salt, but are not
strictly correct.

General Hazards Copper(II) salts are HARMFUL if swallowed. See Hazcard 27A, B & C.

Formula: CuSO
4
.5H
2
-1
Concentration
required
label
100 250 1000
0.1 M 2.50 6.24 24.97 -
0.5 M 12.49 31.21 124.84 -
1.0 M 24.97 62.42 249.68 HARMFUL
Saturated (20 C) 40 100 400 HARMFUL

Formula: CuCl
2
.2H
2
-1
Concentration
required
label
100 250 1000
0.1 M 1.71 4.26 17.05 -
0.5 M 8.52 21.31 85.24 -
1.0 M 17.05 42.62 170.48 (HARMFUL)

Formula: Cu(NO
3
)
2
.3H
2
-1
Concentration
required
label
100 250 1000
0.1 M 2.42 6.03 24.15 -
0.5 M 12.08 30.19 120.75 -
1.0 M 24.15 60.34 241.50 (HARMFUL)
Procedure
Measure out the indicated quantity of copper(II) salt.
Add the solid to about two thirds of the final volume of water in a beaker or jug.

CLEAPSS Recipe Book
38 CLEAPSS 2011
32 Crude oil alternative
Real crude oil (and petrol) contain benzene in concentrations greater than 0.1% and must not be used in
school science (COSHH Regulations).
A synthetic mixture can be prepared using mainly aliphatic hydrocarbons to illustrate the principle of
fractional distillation of crude oil in industry.
An alternative name for petroleum spirit is petroleum ether.
On storing for several months, the lighter fractions evaporate away so the procedure should be tested to see
that enough low boiling point fractions can be distilled off.
Further details on the procedure can be found in L195, Safer Chemicals, Safer Reactions and section 13 of
the Handbook.
Standard fractional distillation equipment using a fractionating column is unsuitable as very high
temperatures are required. Use apparatus similar to the diagram below.

General Hazards Petroleum spirit is HIGHLY FLAMMABLE and HARMFUL (see Hazcard 45A).

Preparing 100 ml of synthetic crude oil
Wear eye protection. Do not prepare the mixture near sources of ignition.
Mix together 55 ml of liquid paraffin (medicinal), 20 ml of paraffin oil (kerosene), 11 ml of white spirit,
4 ml of petroleum ether (100-120 C), 4 ml of petroleum ether (80-100 C) and 6 ml of petroleum ether
(60-80 C).
Add a squeeze of black oil paint (eg, Winsor and Newtons Ivory Black) from a tube and stir well.
After adding to a labelled bottle, shake the mixture well. Label the container HIGHLY FLAMMABLE &
HARMFUL.
Always shake the mixture well before use. The paint will separate out if the bottle is left still for some
time.
Note
The recipe is not written in stone. Another possibility, if you have an even lower fraction of petroleum spirit,
is:
Mix together 50 ml of liquid paraffin (medicinal), 20 ml of paraffin oil (kerosene), 10 ml of white spirit,
5 ml of petroleum ether (100-120 C), 5 ml of petroleum ether (80-100 C), 5 ml of petroleum spirit
(60-80 C) and 5 ml of petroleum spirit (40-60 C).

75 x 10 mm test tubes
to collect the fractions
1 ml of water in a
test tube to compare volumes
(optional)
Side-arm boiling tube
Mineral fibre with
6 ml of crude oil
Heat
Block of wood to
hold the test tubes
0 360 C thermometer
CLEAPSS Recipe Book
CLEAPSS 2011 39
33 2,4-Dinitrophenylhydrazine solution
The solid is also known as 2,4-DNP and the solution is called Bradys reagent.
It is used to identify organic compounds with carbonyl groups by producing orange or yellow insoluble
derivatives which, if purified by recrystallisation, give sharp melting points.
Solid 2,4-DNP is supplied moist as there may be an explosion risk if very dry solid is handled. Unopened
bottles of 2,4-DNP already contain 20% to 33% water.
Two recipes for preparing Bradys reagent are given below. The new recipe using phosphoric acid has been
shown to work and is safer.

General Hazards 2,4-DNP is EXPLOSIVE and TOXIC by inhalation, skin contact and if swallowed
(see Hazcard 30). For information about the alcohols see Hazcard 40A. Both
concentrated sulfuric(VI) acid and phosphoric(V) acid are CORROSIVE (see
Hazcards 98A and 72 respectively).

Formula: (NO
2
)
2
C
6
H
3
NHNH
2
-1

Procedure for making about 100 ml of Bradys reagent
Traditional method New method
Wear eye protection, gloves and work in a fume
cupboard.
Stir 2.7 g of 2,4-DNP with 96 ml of methanol.
Cautiously and slowly add 4 ml of concentrated
sulfuric(VI) acid.
Usually the solid dissolves to form a clear
solution with a little stirring. If cloudy, filter.
Wear eye protection and gloves.
Stir 2 g of 2,4-DNP with 50 ml of 85%
phosphoric(V) acid.
Add 50 ml of ethanol.
Label the solution TOXIC and HIGHLY FLAMMABLE.

Procedure for using Bradys reagent
Wear eye protection, and gloves.
Place 0.5 ml (if liquid) or 0.5 g (if solid) of the carbonyl compound to be identified in a test tube.
Add 5 ml of Bradys reagent. [If crystals do not appear then add a little 2 M sulfuric(VI) acid.]
Cool the solution in an ice bath for 15 to 30 minutes.
Filter off the crystals (vacuum filtration is quicker).
Wash the crystals with water and recrystallise them before obtaining a melting point.
CLEAPSS Recipe Book
40 CLEAPSS 2011
34 3,5-Dinitrosalicylic acid
Also called DNSA or DNS.
3,5-Dinitrosalicylic acid is an alternative to Benedicts solution for testing glucose and other reducing sugars.
It can be used either qualitatively (yellow to red) or quantitatively using colorimetry with a green filter
absorbing at 565 nm.
It is more sensitive to glucose than Benedicts solution.

General Hazards 3,5-Dinitrosalicylic acid is HARMFUL by ingestion and irritating to the eyes and skin.

Preparing DNSA reagent
Wear goggles.
Add 1.0 g of 3,5-dinitrosalicylic acid to 50 ml of water and warm gently to dissolve.
Slowly add 30 g sodium potassium tartrate tetrahydrate, (KNaC
4
H
4
O
6
4H
2
O). The mixture thickens
(like custard).
Add 20 ml of 2 M sodium hydroxide solution and dilute the mixture with water to a final volume of
100 ml.
Label the solution IRRITANT.

Experimental procedure
Mix 0.3 ml of the sample with 3 ml of DNSA reagent. Glass vials are ideal for this.
Heat in hot water (from a kettle) for 5 minutes till a yellow colour develops. The samples do not need
to be boiled.
Remove the vial with tongs and allow to cool in a cold water bath.
The colorimeter will need to be set up with the green filter (about 565 nm).
Use reagent with water as a blank.
There are no further colour changes beyond a glucose concentration of 0.04 M, ie 0.72% (w/v).
Therefore, prepare a glucose standard curve with this concentration as the maximum.
Samples which are believed to contain higher amounts of glucose will need to be diluted before
testing.
CLEAPSS Recipe Book
CLEAPSS 2011 41
35 Drosophila food base
The food-base should pour easily when hot but set firmly when cold. It should also remain firm when the
adult flies emerge.

Preparing the food base
Soak 72 g of oatmeal in 120 ml of pure water.
Dissolve 35 g of treacle (black molasses) in 40 ml of pure water.
Boil 6 g of agar in 400 ml of pure water.
The mould inhibitor is made by dissolving 0.1 g of methyl 4-hydroxybenzoate (Nipagin; see
Hazcard 52) in a small volume of 95% (v/v) industrial denatured alcohol.
Mix all the ingredients and heat in a boiling-water bath, with constant stirring, for at least 15 minutes.
Add more water if the mixture does not pour easily.
Using a funnel, pour the mixture into either milk bottles or specimen tubes, making sure that the
medium does not drip down the sides of the vessels.
Autoclave at 15 psi (121 C) for 15 minutes.
Allow the autoclave to cool naturally after sterilisation is complete; otherwise the medium will boil up
and contaminate the walls of the vessels.

CLEAPSS Recipe Book
42 CLEAPSS 2011
36 Electroplating solutions
There are many formulations for plating solutions. Some are hazardous and where they are not covered by a
model (general) risk assessment, either here or elsewhere, a special risk assessment will be needed before
use by teachers / technicians or students. Help can be obtained from CLEAPSS.

Use Recipe for 100 ml of solution Notes
Copper
electroplating
Use 1 M copper(II) sulfate(VI) solution See Hazcard 27C. Label the
solution HARMFUL.
Nickel
electroplating
Dissolve 5 g of ammonium nickel(II)
sulfate(VI) in 80 ml of water and dilute to
100 ml.
See Hazcard 65B. Label the
solution HARMFUL.
Silver
electroplating
Dissolve 1.6 g of silver nitrate(V) and 32 g of
potassium iodide in 100 ml of water. Add 3
drops of concentrated sulfuric(VI) acid. Use a
carbon rod as the anode.
See Hazcards 87 and 98A. Label
the solution IRRITANT.
Zinc electroplating Dissolve 33 g of zinc sulfate(VI)-7-water in
50 ml of water and dilute to 100 ml. Add five
drops of 2 M sulfuric(VI) acid and 2 spatulas
full of boric acid. Boric acid crystals dissolve
more easily than the powder.
This is an old recipe and the word spatula is
used which is not helpful. The solubility of
boric acid is 6 g per 100 ml of water. 6 g is
probably a sensible amount to add.
See Hazcards 14, 98A and 108B.
It would be advisable to label the
solution HARMFUL.

CLEAPSS Recipe Book
CLEAPSS 2011 43
37 Enzymes
This sheet is applicable to amylase, lipase, trypsin, pepsin etc.
Enzyme preparations are available in the form of powders, capsules, tablets or concentrated solutions.
Stock enzymes should be stored in the refrigerator and discarded when out of date or no longer active.
Enzyme activity varies considerably with age, source and storage conditions. It is therefore essential to
perform preliminary trials to check whether the enzyme is working and whether the endpoint is reached in an
appropriate time. Allow time to order fresh stock in case the sample does not yield satisfactory results.
If the concentration required is not specified on the protocol for the activity or suggested by the supplier,
begin with 1% (w/v), test, and adjust accordingly.
It is often preferable to obtain enzymes from a natural source. Purchased enzymes may have a microbial
origin and are sometimes very heat stable or have unusual pH profiles so that investigations of the influence
of temperature or pH on enzyme activity yield unexpected results. For example, saliva is often more reliable
as a source of amylase than commercial products, which may also be contaminated with sugar. Health risks
are minimal if each pupil uses his/her own saliva and rinses out contaminated glassware afterwards, prior to
soaking in sodium chlorate(I) solution before normal washing up. Follow procedures in the Handbook.
Note that for many enzymes, the pH will need to be adjusted to the optimum; check the details provided or
the protocol.
Enzyme powders may, in extreme cases, cause sensitisation. It is best to make up solutions in a fume
cupboard which is not switched on. This helps to protect the operator from breathing in the powder in case it
disperses. An operating fume cupboard may cause the powder to blow around. In case of a spill, the fume
cupboard should be switched on to vent the powder.

General Hazards See Hazcard 33. All enzyme powders are HARMFUL and may cause sensitisation.
Any solutions greater than 1% should be considered as IRRITANT.

Preparing 100 ml solution of 1% enzyme solution
Wear eye protection and disposable nitrile gloves when making up solutions from the powders.
Avoid inhaling the powder. It may be wise to carry out the weighing and preparation in a fume
cupboard (with the fan switched off to avoid the draught affecting the balance).
Measure out 1 g of the enzyme.
Add the solid to about 70 ml of water or buffer (if needed for the activity) in a beaker.
Stir to dissolve (do not warm the solution).
Pour the solution into an appropriate measuring cylinder/volumetric flask.
Dilute to the final volume with pure water.
Pour into a labelled bottle and mix well. Store in the refrigerator or on ice during use.
Carry out the procedure that the students will undertake and consult with the teacher to confirm that
the results are satisfactory. It may be necessary either to dilute the solution further with more water or
add more of the enzyme.

Sodium tauroglycocholate (Bile salts)
General Hazards See Hazcard 95C. Although low hazard, bile salts have an unpleasant odour and
may be better prepared in the fume cupboard.

Preparing 100 ml of 3% bile salt solution
Weigh out 3 g of bile salts.
Dissolve the salt in 70 ml of water.
Make up to 100 ml.

CLEAPSS Recipe Book
44 CLEAPSS 2011
38 Etching solutions
There are many formulations for etchants. Some are hazardous and where they are not covered by a model
(general) risk assessment, here or elsewhere, a special risk assessment will be needed before use by
teachers / technicians or students. Help can be obtained from CLEAPSS.

Use Recipe for 100 ml of solution Notes
Aluminium
etching
Use 2 M sodium hydroxide solution at 60-70 C. See Hazcard 91, Wear goggles.
Copper etching Dissolve 20 g of iron(III) chloride-6-water in
80 ml of water and 20 ml of concentrated
hydrochloric acid (36% w/v).
See Hazcards 47A and 55C.
Label the solution IRRITANT.
Iron etching 1
(Frys reagent)
Dissolve 59 g of copper(II) choride-2-water in
40 ml of water and add 60 ml of concentrated
hydrochloric acid (36% w/v).
See Hazcards 27A and 47A. Label
the solution CORROSIVE.
Iron etching 2
(Nital solution)
Add 2 ml of concentrated nitric(V) acid to 98 ml
of ethanol. Under no circumstances use more
than 2 ml of nitric(V) acid in making up 100 ml
of solution.
See Hazcards 40A and 67. Label
the solution HIGHLY FLAMMABLE &
HARMFUL. Do not store.
Lead etching Wear goggles and chemical resistant gloves.
Before use, mix together 50 ml of 5 M nitric(V)
acid and 50 ml of 15% (w/v) ammonium
molybdate(VI) solution. Apply with a swab for
30 s before rinsing.
See Hazcards 9A and 67. Label
the solution CORROSIVE.

CLEAPSS Recipe Book
CLEAPSS 2011 45
39 Ethanoic acid
Also known as glacial acetic acid.
The concentration of this solution is 17.4 mol dm
-3
.
On cold days, the acid freezes to form a glacial solid. In this case, place the bottle in warm water for several
hours before making the diluted sample, but make sure the label does not come off.

Formula: CH
3
COOH Molar mass: 60.05 g mol
-1

General Hazards See Hazcard 38A. Ethanoic acid is CORROSIVE. It has a very sharp odour.

Volume (ml) of concentrated acid to be used
Concentration
required
label
500 1000 2500
0.1 M Ten-fold dilution of 1 M solution -
0.5 M 14 29 71 -
1 M 29 57 143 -
2 M 57 114 286 IRRITANT

Procedure
Wear goggles (a face shield is preferable for large volumes) and chemical resistant gloves.
Use the fume cupboard.
Measure out the indicated volume of ethanoic acid in a measuring cylinder.
Add the liquid to about two thirds of the final volume of water in a beaker or laboratory jug.
Stir the mixture well.
CLEAPSS Recipe Book
46 CLEAPSS 2011
40 Fehlings solutions
Benedicts solution or DNSA are safer alternatives when testing for sugars.
Fehling's solution can only be used to test for aliphatic aldehydes, whereas Tollen's reagent (see Recipe
Sheet 102) can be used to test for both aliphatic and aromatic aldehydes.
Fehling's solutions A & B need to be made and stored separately. They are mixed just prior to use.
Sandells solution (which is little known and not cited in exam school texts; see Recipe Sheet 76) can be
made and stored for long periods. The concentration of sodium hydroxide is only 0.4 M in the solution
(Fehlings solution B is close to 4M).

General Hazards Copper(II) sulfate(VI) crystals are HARMFUL if swallowed (see Hazcard 27C). 2M
sulfuric(VI) acid, solid sodium hydroxide and 0.5 M sodium hydroxide solution are
CORROSIVE (see Hazcards 91 and 98A).

Preparing 100 ml of Fehlings solution
Fehlings solution A
Measure out 7 g of copper(II) sulfate(VI)-5-water and dissolve this in 60 ml of water.
If the solution appears cloudy add 10 ml of 2 M sulphuric acid.
Make up to 100 ml with water. Label the bottle.

Fehlings solution B
Wear goggles.
Measure out 15.4 g of sodium hydroxide and 35 g of potassium sodium tartrate.
Add the solids to 60 ml of water on a magnetic stirrer. Make up to 100 ml with water. Label the bottle
and add a CORROSIVE warning.

Solutions A and B should be mixed in equal volumes, just prior to use. During the test, do not heat directly
over a Bunsen burner: use a boiling water bath.

CLEAPSS Recipe Book
CLEAPSS 2011 47
41 Fixatives used before preserving biological specimens
Fixatives kill cells whilst retaining their structure.
Although safer preservatives are available as alternatives to methanal, it is necessary to use methanal in
many fixatives. (See Recipe Sheet 74 for preservatives.)
More information about the procedures for fixing and preserving specimens can be found in the Laboratory
Handbook, section 15.

Methanal solution
for fixing animal
tissue
Dilute 10 ml of 40% methanal solution to
100 ml. Leave specimen in this solution
for 16-48 hours before preserving.
For methanal, see Hazcard 63. Label
the final solution HARMFUL. Use a
fume cupboard, wear eye protection
and disposable nitrile gloves.
Formalin-alcohol
fixative for plant
tissue
Mix together 90 ml of ethanol with 10 ml
of 40% methanal solution. Leave
specimen in this solution for 12 hours
before preserving.
For ethanol; see Hazcard 40A. For
methanal, see Hazcard 63. Label the
solution HIGHLY FLAMMABLE and
HARMFUL. Use a fume cupboard,
wear eye protection and disposable
nitrile gloves.
Formalin-aceto-
alcohol (FAA)
fixative for plant
tissue
Mix together 5 ml of 40% methanal and
5 ml of glacial ethanoic acid in 90 ml of
70% ethanol. Leave specimen in this
solution for 12 hours before preserving.
methanal, see Hazcard 63. For
ethanoic acid, see Hazcard 38.
Label the solution HIGHLY FLAMMABLE
& HARMFUL. Use a fume cupboard,
wear eye protection and disposable
nitrile gloves.
Acetic alcohol
(Clarkes fluid) (a
cytological fixative
especially for
chromosomes)
Mix 25 ml of glacial ethanoic acid with
75 ml of absolute alcohol (Industrial
denatured alcohol is normally
adequate).
Label the solution HIGHLY FLAMMABLE
and CORROSIVE. Use a fume
cupboard, wear eye protection and
disposable nitrile gloves.

CLEAPSS Recipe Book
48 CLEAPSS 2011
42 Gases less commonly used in schools

Gas Hazard
information
Minimum amount of reagents to prepare 1000 ml of gas
Carbon
monoxide
See Hazcard 21 Concentrated sulfuric(VI) acid is slowly added to 2.9 g of solid sodium
methanoate. Collect over water.
Dinitrogen
monoxide
See Hazcard 68 5.8 g of hydroxyammonium chloride in 30 ml of water is added to a warm
solution containing 81 g of ammonium iron(III) sulfate(VI) in 160 ml of
water. Collect over warm water.
Hydrogen
chloride
See Hazcard 49 2.1 ml of concentrated sulfuric(VI) acid is slowly added to excess solid
sodium chloride. Collect by downward delivery (a tube into the bottom of
an upright gas jar).
Hydrogen
sulfide
See Hazcard 51 42 ml of 2 M hydrochloric acid is added to excess iron(II) sulfide. Collect
over hot water.
Nitrogen See Hazcard 93 2.9 g of sodium nitrate(III) (nitrite) dissolved in a little water is warmed
gently with excess ammonium chloride. Pass the gas through water to
remove any oxides of nitrogen formed as a side reaction, and then collect.
Nitrogen
dioxide
See Hazcard 68 8 ml of concentrated nitric(V) acid (70%) is added to an excess of copper
turnings. Collect by downward delivery (a tube into the bottom of an
upright gas jar).
Nitrogen
monoxide
See Hazcard 68 30 ml of half-concentrated nitric(V) acid (35%) is added to an excess of
copper turnings. Collect over water.
OR
2.9 g of sodium nitrate(III) (nitrite) dissolved in 6 ml of water is added to
12 g of hydrated iron(II) sulfate(VI) and enough 5 M hydrochloric acid is
added to cover the crystals. Collect over water.

CLEAPSS Recipe Book
CLEAPSS 2011 49
43 Hydrochloric acid
Hydrochloric acid is available from all major educational suppliers as 35 - 38% (w/w) solution with a specific
gravity of 1.18. If you wish to know the dilution factors for other commercial concentrations (eg, 32%), then
contact CLEAPSS.
For accurate work, the concentration should always be found by titrating against a standard solution of
sodium carbonate.
The concentration of concentrated hydrochloric aid is about 11.7 mol dm
-3
.
If kept for long periods, the concentration of 35 - 38% (w/w) hydrochloric acid decreases as the gas diffuses
into the atmosphere.
Do not make dilute solutions for the first time without seeking practical advice from a more experienced
colleague.

Formula: HCl Density: 1.18 g cm
-3
-1

General Hazards See Hazcard 47A. Hydrochloric acid is CORROSIVE. It has a very sharp odour.

Concentration
required
label
500 1000 2500
0.4 M 17 34 84 -
0.5 M 21 42 105 -
1 M 42 84 209 -
2 M 84 167 417 IRRITANT

Procedure
Wear goggles (a face shield is preferable when handling large volumes) and chemical resistant
gloves.
Use a fume cupboard. Take care opening a bottle on a hot day.
Measure out the indicated quantity of concentrated hydrochloric acid in a measuring cylinder.

CLEAPSS Recipe Book
50 CLEAPSS 2011
44 Hydrogen gas
General Hazards See Hazcards 27C, 48 & 98. Hydrogen is extremely flammable. There must be no
Bunsen burner being used within 1 m of the apparatus.
Note that the first gas collected consists of air and hydrogen. This is an explosive
mixture and should be discarded.

Wear goggles.
Place several granules of zinc in the
generator.
The reaction needs to be speeded up at the
start, to flush out air quickly.
Add about 5 ml of 1 M copper sulfate(VI)
(HARMFUL) down the thistle funnel. This reacts
with zinc, coating it with copper which acts as
a catalyst.
Add 2 M sulfuric(VI) acid (CORROSIVE) down
the thistle funnel.
Collect the gas in the 250 ml measuring
cylinder. Once the measuring cylinder is full,
its contents (largely air) must be discarded.
Now gas jars, boiling tubes, syringes and soap
bubbles of pure hydrogen can be prepared.

Dry hydrogen [for burning or reducing copper(II) oxide]
Use a U-tube filled loosely with calcium
chloride granules (not powder) and collect the
first gases in measuring cylinder one size
larger than the hydrogen generator.
Once the measuring cylinder is full, disconnect
at A, and attach other equipment but before
using the hydrogen, leave time to flush the
air out of this newly-attached equipment.
The gas collected in the measuring cylinder
can be discarded.

250 ml measuring
cylinder
Zinc
Hydrogen
generator
250 ml
plastic bottle
1 to 5 ml of 1M
copper(II) sulfate(VI) plus
2M sulfuric acid
A
500ml measuring
cylinder;
to ensure
all air is flushed
through
U-tube
Calcium chloride
CLEAPSS Recipe Book
CLEAPSS 2011 51
45 Hydrogen peroxide
In the UK, the concentration of hydrogen peroxide is given in terms of the volume of oxygen it can produce;
thus 1 ml of 10 vol hydrogen peroxide is capable of producing 10 ml of oxygen gas at 0 C and 1atm.
The maximum concentration of purchased hydrogen peroxide is 100 vol and even this solution will become
more dilute with time.
The concentration of 100 vol solution is 8.3 mol dm
-3
.
Commercial hydrogen peroxide solutions contain an inhibitor to slow down decomposition. During the
preparation of diluted solutions, the inhibitor is diluted so solutions will deteriorate quite quickly. They should
be prepared when required and not stored for long periods.
For accurate work you will need to standardise the solution against a known solution of potassium
manganate(VII).
For activities with catalase, start with a 1 M solution and adjust subsequent concentrations depending on
observed results.

Formula: H
2
O
2
-1

General Hazards See Hazcard 50. 100 vol hydrogen peroxide is HARMFUL.

Concentration
required
Volume (ml) of solution required Hazard
warning
label
250 1000 2500
0.08 M 1 vol 0.29% Twenty-fold dilution of the 20 vol solution -
0.1 M 1.2 vol 0.34% Ten-fold dilution of the 1 M solution -
0.34 M 4 vol 1.14% Five-fold dilution of the 20 vol solution -
1 M 12 vol 3.3% 29 115 288 -
1.7 M 20 vol 5.7% 50 200 500 IRRITANT
3.4 M 40 vol 11.4% 100 400 1000 IRRITANT

Procedure
Wear eye protection and gloves.
Measure out the indicated quantity of 100 vol hydrogen peroxide in a measuring cylinder.
Add the liquid to about two thirds of the final volume of water in a beaker or laboratory jug and stir.
CLEAPSS Recipe Book
52 CLEAPSS 2011
46 Indicators (acid-base)
General Hazards See Hazcard 32. The hazards of many dyes and indicators are not well known.
Many are made up in ethanol which is HIGHLY FLAMMABLE & HARMFUL (see
Hazcard 40A).

Indicator Quantity
(g)
Volume of
IDA
Acid Alkali pH range
Bromophenol blue 0.4 200 Yellow Blue 3.0 - 4.5
Methyl orange 0.4 200 Red Yellow 3.0 - 4.5
Bromocresol green 0.4 200 Yellow Blue 4.0 - 5.4
Methyl red 0.2 300 Red Yellow 4.0 - 6.0
Bromothymol blue 0.4 200 Yellow Blue 6.0 - 7.5
Phenol red 0.4 200 Yellow Red 6.6 - 8.0
Neutral Red 0.4 200 Red Yellow 6.8 - 8.0
Cresol red 0.2 200 Yellow Purple 7.2 - 8.8
Thymol blue 0.4 200 Yellow Violet 8.0 - 9.6
Phenolphthalein 1.0 600 Colourless Mauve 8.0 - 10.0
Thymolphthalein 2.0 1000 Colourless Blue 9.3 - 10.5

Procedure to prepare 1000 ml of indicator solution
Wear eye protection and use a fume cupboard when dispensing from bottles of solid indicators to
avoid raising dust. Use gloves to avoid staining the skin.
Dissolve the quantity of indicator in the volume of industrial denatured alcohol (IDA) given in the table
in a suitable beaker.
Transfer solution to a 1000 ml measuring cylinder and make up to 1000 ml with water.

Litmus solution
Dissolve 1 g of litmus in 1000 ml of water. Other recipes suggest up to 10 g of litmus but this may
depend upon the age of the sample. Filter if necessary.

Home-made indicators from fruits and vegetables
Coloured aqueous solutions can be made from red cabbage, flower petals, blackberries, beetroot, etc, frozen
in ice-cube trays and stored for months.

Screened methyl orange
The normal colour change of some indicators is not always clear and may be altered by screening. The
most common screened indicator used in schools is screened methyl orange.
Dissolve 1 g of methyl orange and 2.6 g of xylene cyanol FF in 1000 ml of water. The end point is a grey
colour.
CLEAPSS Recipe Book
CLEAPSS 2011 53
47 Indicator (universal)
Ethanol is HIGHLY FLAMMABLE & HARMFUL (see Hazcard 40A).

Procedure to prepare 1000 ml of Yamada Universal indicator solution
Wear eye protection and use a fume cupboard (not switched on) to avoid raising dust when
dispensing from bottles of solid indicators. Use gloves to avoid staining the skin.
Place 0.25 g of bromothymol blue, 0.025 g of thymol blue, 0.063 g of methyl red and 0.5 g of
phenolphthalein in an appropriate beaker.
Add 500 ml of ethanol and stir to dissolve the dyes.
Add 500 ml of pure water.
Pour into a labelled 1000 ml bottle and mix well.
Label this solution (which is red) FLAMMABLE.
The colours of this solution when added to buffers are shown below.

3 3.5 4 4.5 5 5.5 6 6.5 7 7.5 8 8.5 9 9.5 10 10.5 11
CLEAPSS Recipe Book
54 CLEAPSS 2011
48 Indicators (carbon dioxide)

Many are made up in ethanol which is HIGHLY FLAMMABLE & HARMFUL (see
Hazcard 40A).

Hydrogencarbonate (bicarbonate) indicator: Preparation of the stock solution:
Use a fume cupboard (not switched on) to avoid raising dust when dispensing from bottles of solid
indicators. Use gloves to avoid staining the skin.
Dissolve 0.20 g of thymol blue and 0.1 g of cresol red in 20 ml of ethanol.
Dissolve 0.85 g of sodium hydrogencarbonate in about 200 ml of freshly-boiled distilled water.
Add the ethanolic dye solution and dilute to 1000 ml with water.

For use:
Dilute the stock solution ten times with freshly-boiled distilled water.
Bubble air through the diluted solution to equilibrate it with atmospheric carbon dioxide.
When ready for use, the solution should be a deep cherry red colour in a bottle (in a test tube the
colour of the solution is less intense). If difficulties are experienced (eg, if distilled water is too acidic),
try adding a pinch more sodium hydrogencarbonate. Avoid breathing over open vessels of the diluted
indicator; the carbon dioxide exhaled may alter its pH.

Bromothymol blue indicator
This can be used as an alternative to the above indicator in carbon dioxide uptake/release studies in biology.
Some authorities report fewer difficulties in obtaining good results. The colour change is to yellow (more
acid) and blue (more alkaline).

Use a fume cupboard (not switched on) to avoid raising dust when dispensing from bottles of solid
indicators. Use gloves to avoid staining the skin.
Dissolve 0.02 g of bromothymol blue in 20 ml ethanol before diluting to 1 litre with water.

CLEAPSS Recipe Book
CLEAPSS 2011 55
49 Indicators for redox, precipitation and complexometric titrations
Cerium(IV) titrations
Dissolve 1.49 g of 1:10-phenanthroline in 100 ml of water and add
0.70 g of iron(II) sulfate(VI). (Ferroin indicator).
Wear eye protection. 1:10-
phenanthroline is TOXIC. The solution is
low hazard.
Dissolve 0.1 g of N-phenylanthranilic acid in 5 ml of
0.1 M sodium hydroxide. Dilute to 100 ml with water.
See Hazcard 78.
Dichromate(VI) titrations
Dissolve 0.1 g of N-phenylanthranilic acid in 5 ml of
0.1 M sodium hydroxide. Dilute to 100 ml with water.
See Hazcard 78.
Dissolve 0.2 g of sodium or barium diphenylamine-4-sulfonate
[diphenylamine-4-sulfonic acid (sodium salt)] in 100 ml of water.
See Hazcard 10B. Sodium
diphenylamine-4-sulfonate is an
IRRITANT. Wear eye protection.
Starch indicator for thiosulfate/iodine titrations (and detecting iodine)
Mix 1 g of soluble starch with a small amount of pure water and
stir it vigorously to form a paste.
Boil 100 ml of pure water and add it to the paste with stirring and
allow to cool.
This solution (it is really a suspension)
does not keep. Dispose of it if it starts to
go cloudier or lumpy.
Silver nitrate(V) titrations
Dissolve 5 g of potassium chromate(VI) in 100 ml of pure water. See Hazcard 78. Wear goggles. Label
this solution TOXIC.
Dissolve 0.1 g of eosin in a mixture of 70 ml of ethanol and
30 ml of water.
See Hazcard 32 & 40A. Label this
solution HIGHLY FLAMMABLE & HARMFUL.
Precipitate becomes pink.
Dissolve 0.1 g of fluorescein (sodium salt) in 100 ml of water. See Hazcard 32 & 40A. Label this
solution HARMFUL. Precipitate becomes
pink.
(For Volhards method.) Dissolve 10 g of ammonium iron(III)
sulfate(VI).12-water in 90 ml of water and add enough 2 M
nitric acid to clear the solution and prevent hydrolysis. Make up to
100 ml. Use 1 ml per titration.
See Hazcard 55C. Colour change: white
ppt to orange red colouration.
EDTA titrations
Dissolve 1 g of the Eriochrome black T (Solochrome black) in
100 ml of ethanol or grind 1 g of the solid dye with 100 g of
sodium chloride. Use about 0.2 g of this solid mixture for each
titration.
solution HARMFUL. (Colour change is red
to blue.)
Grind 1 g of murexide with 100 g of sodium chloride. Use 0.2 g for
each titration.
solution HARMFUL. (Colour change is red
to blue.)

CLEAPSS Recipe Book
56 CLEAPSS 2011
50 Iodine solution
Iodine is only sparingly soluble in water (0.3 g/L).
It is usual to dissolve it in aqueous potassium iodide solution (KI) or organic solvents such as ethanol.
The procedure will take time even with stirring.
A 0.01 M solution is suitable as a test reagent for starch.
The concentration of solutions decreases with storage. Check that the solutions work before use in the
laboratory.

Formula: I
2
-1

General Hazards See Hazcard 54. The organic solvents used are also hazardous and the relevant
Hazcard should be consulted.

Concentration
required
warning
label
100 250 1000
0.1 M 8 g of KI + 2.54 g of I
2
24 g of KI + 6.35 g of I
2
80 g of KI + 25.38 g of I
2
-

Procedure
Wear eye protection and work in a well-ventilated room. Wear disposable nitrile gloves.
Measure out the indicated quantity of potassium iodide (KI) into an appropriate beaker.
Moisten the potassium iodide with a few drops of water.
Measure out the indicated quantity of iodine and add it to the moistened potassium iodide.
Add a small volume of water and stir. When no more iodine appears to dissolve, add some more water
and stir. Keep repeating this procedure until all the iodine has dissolved.
Pour the solution into a measuring cylinder and dilute to the final volume. Make sure there are no bits
of iodine remaining. If there are, return the solution to the beaker and leave it on a magnetic stirrer for
several minutes.
Add the solution to a labelled bottle and mix well.

CLEAPSS Recipe Book
CLEAPSS 2011 57
51 Iron(II) solutions
Comments about iron(II) salts
Iron(II) sulfate(VI)-7-water crystals are green but they sometimes turn white through loss of water of
crystallisation (efflorescence) or brown through oxidation during storage.
Iron(II) chloride-4-water should also be green but it is even more susceptible to oxidation than iron
sulfate(VI), changing to a brown solid. It is often quoted in experimental worksheets for displacement
reactions but iron(II) sulfate(VI) or diammonium iron(II) sulfate(VI) work just as well.
Neutral solutions of iron(II) salts turn brown very quickly on standing because of oxidation by air.
Iron(II) salts are best prepared in an acidic solution rather than water. It is safer to use this procedure than to
start adding concentrated sulfuric(VI) acid at the end, as is often stated in older recipe books.
Diammonium iron(II) sulfate(VI) is more stable to air during storage. It is the preferred salt for titration
experiments (see Recipe sheet 52).
Do not to store these acidic iron(II) solutions for long periods. A clear green colour gradually turns to dirty
green because of air oxidation.

General Hazards See Hazcards 55B and 98A. Label the final solution IRRITANT because of the acid
present.
Iron(II) sulfate(VI)-7-water
Also known as ferrous sulfate heptahydrate.
Formula: FeSO
4
.7H
2
-1

Concentration
required
label
100 250 1000
0.01 M Ten-fold dilution of a 0.1 M solution with 0.1 M sulfuric(VI) acid -
0.1 M 2.78 6.95 27.80 IRRITANT
1 M 27.80 69.50 278.01 IRRITANT

Iron(II) chloride-4-water
Do not use the anhydrous salt.
Formula: FeCl
2
.4H
2
-1

Concentration
required
label
100 250 1000
0.01 M Ten-fold dilution of a 0.1 M solution with 0.1 M hydrochloric acid -
0.1 M 1.99 4.97 19.88 IRRITANT
1 M 19.88 49.7 198.80 IRRITANT

Procedure
Measure out the indicated quantity of iron(II) salt.
Add the solid to about two thirds of the final volume of 1 M sulfuric(VI) acid for the sulfate(VI) or 1 M
hydrochloric acid for the chloride in a beaker or laboratory jug.
either 1 M sulfuric(VI) acid for the sulfate(VI) or 1 M hydrochloric acid for the chloride to the required
level.
CLEAPSS Recipe Book
58 CLEAPSS 2011
Diammonium iron(II) sulfate(VI)
Also known as ammonium ferrous sulfate(VI) or Mohrs salt.

Formula: (NH
4
)
2
SO
4
.FeSO
4
.6H
2
-1

General Hazards 1 M sulfuric(VI) acid; Hazcard 98A. Iron(II) salts; Hazcard 55B.

Concentration
required
label
100 250 1000
0.01 M Ten-fold dilution of a 0.1 M solution with 0.1 M sulfuric(VI) acid IRRITANT
0.1 M 3.92 9.80 39.21 IRRITANT
0.5 M 19.61 49.02 196.07 IRRITANT

Procedure
Measure out the indicated quantity of diammonium iron(II) sulfate(VI).
Add the solid to about two thirds of the final volume of 1 M sulfuric(VI) acid in a beaker or laboratory
jug.
1 M sulfuric(VI) acid to the required level.
Pour into a labelled bottle and mix well. Add hazard warning.
CLEAPSS Recipe Book
CLEAPSS 2011 59
52 Iron (III) solutions
Aqueous solutions of iron(III) salts do not keep if made up solely in water. If required for a longer period, an
acid should be added. Even in acid solution, there is gradual deterioration and a solid often separates out on
storage.
Iron(III) chloride (also known as ferric chloride) is available in two forms, hydrated iron(III) chloride and
anhydrous iron(III) chloride. The anhydrous solid should not be used to make iron(III) solutions. It does not
dissolve but reacts exothermically with water, to form iron(III) hydroxide and hydrogen chloride gas.
Iron(III) chloride-6-water does not store well as it absorbs water (hygroscopic).
Iron(III) sulfate(VI) has a variable amount of water of crystallisation (9-water is just one of the varieties on
sale). It does not dissolve completely and the solution will need to be filtered (or the solid allowed to settle)
before use.
Ammonium iron(III) sulfate(VI) is much easier to use and is suitable for most applications. The lilac salt
stores quite well but the surface sometimes becomes brown/yellow if the solid has become damp. The
brown/yellow solid should be removed before weighing. The solid also deteriorates if exposed to heat and
light. The solution is brown, Solutions should not be heated to accelerate dissolving as colloidal solutions can
form.
See the etching solution on Recipe sheet 38 if the iron(III) chloride is to be used for this purpose.
Iron (III) chloride
Also known as ferric chloride.

Formula: FeCl
3
.6H
2
-1

General Hazards See Hazcard 55C. Do make sure you are using then hydrated salt and not the
anhydrous salt.

Concentration
required
label
100 250 1000
0.01 M Ten-fold dilution of a 0.1 M solution with 0.2 M hydrochloric acid -
0.1 M 2.70 6.76 27.03 IRRITANT
1 M 27.03 67.58 270.30 IRRITANT

Procedure
Measure out the indicated quantity of iron(III) chloride-6-water.
Add the solid to about two thirds of the final volume of 1 M hydrochloric acid in a beaker or laboratory
jug.
Pour the solution from the beaker into an appropriate measuring cylinder or a laboratory jug and add
1 M hydrochloric acid to the required level.

CLEAPSS Recipe Book
60 CLEAPSS 2011
Iron(III) sulfate(VI)

Formula: Fe
2
(SO
4
)
2
.9.H
2
-1

General Hazards See Hazcards 55C and 98. 1 M sulfuric(VI) acid. Label the final solution IRRITANT
because of the acid present.

Concentration
required
label
100 250 1000
0.1 M 5.6 14.1 56.2 IRRITANT
0.5 M 28.1 70.3 281 IRRITANT
1 M 56.2 140.5 562 IRRITANT

Ammonium iron(III) sulfate(VI)

Formula: NH
4
Fe(SO
4
)
2
.12.H
2
-1

General Hazards See Hazcards 55C and 98. 1 M sulfuric(VI) acid. Label the final solution IRRITANT
because of the acid present.

Concentration
required
label
100 250 1000
0.1 M 4.82 12.05 48.22 IRRITANT
0.5 M 24.11 96.4 241.10 IRRITANT

Procedure
Measure out the indicated quantity of the iron(III) salt.
Add the solid to about two thirds of the final volume of 1 M sulfuric(VI) acid in a beaker or laboratory
jug.
1 M sulfuric(VI) acid to the required level.

Ammonium iron(III) citrate and blueprints
Wear eye protection. Dissolve 3 g of iron(III) chloride-6-water and 3 g of citric acid in 50 ml of water in a
250 ml plastic beaker. Add about 4.5 g of ammonium carbonate in small amounts, stirring each time. There
is quite a lot of frothing and the solution turns green. This solution will keep.
Blue print solution: Add 5 g of potassium hexacyanoferrate(III) and stir. This addition has to be made just
before it is required.
Procedure
The solution is painted onto paper in dull indoor light (preferably in a cupboard) and hung to dry over paper
towels or blotting paper. An opaque object (eg, scissors) is placed on the dry paper and left in the light for a
day or two although, on a sunny day, 20 minutes may well be enough for light to convert soluble iron(III) salts
to insoluble iron(II) salts.
Wearing gloves, rinse the paper in a bowl of water to remove the solution leaving a colourless image on a
Prussian Blue background. This can be messy and may stain clothing and unprotected hands. The paper is
again hung to dry over absorbent paper.
CLEAPSS Recipe Book
CLEAPSS 2011 61
53 Lead(II) nitrate(V)
Tap water contains sulfate and carbonate ions which will precipitate out as insoluble lead compounds if lead
ions are added. Lead salts should, thereore, be prepared in distilled or deionised water.
Lead(II) nitrate(V) is used in preference to lead(II) ethanoate (acetate) as it appears to be slightly less
hazardous.
Disposal issues: see section 7.5 of the Handbook.

Formula: Pb(NO
3
)
2
-1

General Hazards See Hazcards 57A & B.

Concentration
required
label
100 250 1000
0.005 M Twenty-fold dilution of a 0.1 M solution with water
or ten-fold dilution of 0.05 M solution
-
0.01 M Ten-fold dilution of a 0.1 M solution with water TOXIC
0.05 M 1.66 16.55 TOXIC
0.1 M 3.31 8.28 33.12 TOXIC
1 M 33.12 82.80 331.21 TOXIC
Saturated (20 C) 56 140 560 TOXIC

Procedure
Wear goggles and disposable nitrile gloves. If necessary, weigh materials on a balance placed in a
fume cupboard which is not switched on and the sash pulled down.
Measure out the indicated quantity of lead(II) nitrate(V).
CLEAPSS Recipe Book
62 CLEAPSS 2011
54 Lithium chloride
Formula: LiCl Molar mass: 42.39 g mol
-1

General Hazards Lithium salts are HARMFUL if swallowed and irritants. See Hazcards 47A and 58.


Concentration
required
label
100 250 1000
0.5 M 2.12 5.30 21.20 -
1.0 M 4.24 10.60 42.39 -

Procedure
Measure out the indicated quantity of lithium chloride.
Pour into a labelled bottle. Add a hazard warning if appropriate.

CLEAPSS Recipe Book
CLEAPSS 2011 63
55 Magnesium sulfate(VI)
Magnesium sulfate(VI)-7-water loses water of crystallisation slowly on standing to the air and becomes
powdery (efflorescent). However, it is more convenient to make solutions from the hydrated salt than the
anhydrous salt which is also available from suppliers.
The anhydrous salt readily absorbs water from the atmosphere during storage.

Formula: MgSO
4
.7H
2
-1

General Hazards See Hazcard 59B.

Concentration
required
label
100 250 1000
0.1 M 2.47 6.16 24.65 -
0.5 M 12.32 30.81 123.24 -
1.0 M 24.65 61.62 246.47 -
Saturated (20 C) 75 190 750 -

Procedure
Measure out the indicated quantity of hydrated magnesium sulfate(VI).
CLEAPSS Recipe Book
64 CLEAPSS 2011
56 Manganese(II) sulfate(VI)
Manganese(II) sulfate(VI) forms many hydrates. The 4 water and 1 water varieties appear to be the most
readily available.
The values in brackets in the table below are for the monohydrate.

Formula: MnSO
4
.4H
2
-1
Formula: MnSO
4
.H
2
-1

General Hazards Manganese(II) sulfate(VI) is HARMFUL. See Hazcard 60.

Concentration
required
label
100 250 1000
0.1 M 2.23 (1.69) 5.58 (4.23) 22.31 (16.90) -
0.5 M 11.15 (8.45) 27.89 (21.13) 111.54 (84.51) -
1.0 M 22.31 (16.90) 55.77 (42.26) 223.07 (169.02) HARMFUL
Saturated (20 C) 100 (45) 250 (115) 1000 (450) HARMFUL

Procedure
Measure out the indicated quantity of hydrated manganese(II) sulfate(VI).
Pour into a labelled bottle. Add a hazard label if appropriate.
CLEAPSS Recipe Book
CLEAPSS 2011 65
57 Mercury solutions
Only make what is needed. Diluting to 0.04 M reduces the level of hazard to TOXIC.
The main issue with mercury salts is that they should not be put down the drain. The waste should be
collected, stored as toxic waste and removed to sealed landfill by a registered hazardous waste collector.
Although labelled toxic and having a notorious history, using an inorganic salt will not kill you. However, the
risk assessment for the activity must establish that the use of the mercury compound is really necessary and
that no alternative is possible.

General Hazards Mercury salts are VERY TOXIC. See Hazcards 62 & 67.

Mercury(I) nitrate [also known as dimercury(I) nitrate(V)]

Make up in dilute nitric(V) acid to produce a clear solution.

Formula: Hg
2
(NO
3
)
2
.2H
2
-1

Procedure for preparing 100 ml of 0.02 M of dimercury(I) nitrate(V)-2-water solution [which is 0.04 M
with respect to the mercury(I) ion]
Wear goggles.
Dissolve 1.12 g of the solid in 70 ml of 1 M nitric(V) acid.
Make up to 100 ml with 1 M nitric(V) acid.
Label the solution CORROSIVE and TOXIC.

Mercury(II) chloride

Formula: HgCl
2
-1

Procedure for preparing 100 ml of 0.04 M of mercury(II) chloride solution
Wear goggles.
Dissolve 1.09 g of mercury(II) chloride-2-water in 70 ml of water.
Label the solution TOXIC.
CLEAPSS Recipe Book
66 CLEAPSS 2011
58 Methanal solution
Also known as formalin or formaldehyde solution.
The commercial solution is a 40% (w/v) solution of methanal in water with stabiliser added.
Methanal is a gas at room temperature but it dissolves in water. If the bottle top is not secure then it will
gradually lose gas and its concentration will fall.
The concentration of the 40% solution is 11.3 mol dm
-3
.

Formula: HCHO Molar mass: 30.03 g mol
-1

General Hazards See Hazcard 63. It can cause burns, conjunctivitis or sensitisation by skin contact.

Volume of liquid required
Concentration
required
label
500 1000 2500
0.3% 0.1 M Ten-fold dilution of a 0.1 M solution with water IRRITANT
1% 0.3 M 13 25 63 HARMFUL
3% 1 M 38 75 190 HARMFUL
4% 1.3 M 50 100 250 HARMFUL

Procedure
Use a fume cupboard to handle the concentrated solution; wear goggles (a face shield is preferable
when handling large volumes) and wear disposable nitrile gloves.
Measure out the indicated volume of 40% methanal solution in a measuring cylinder.
Add the liquid to about two thirds of the final volume of water in a suitable beaker or laboratory jug and
stir.
Pour into a labelled bottle and mix well. Add the appropriate hazard warning.
CLEAPSS Recipe Book
CLEAPSS 2011 67
59 Methanoic acid
Also known as formic acid.
It is usually supplied as a 90% (w/w) solution.
-3
.

Formula: HCOOH Density: 1.22 g cm
-3
-1

General Hazards See Hazcard 38A. Methanoic acid is CORROSIVE. It has a sharp odour.

Concentration
required
warning label
250 500 1000 2500
1 M 10.5 21 42 105 -
2 M 21 42 84 210 IRRITANT

Procedure
Wear goggles (a face shield is preferable for large volumes) and chemical-resistant gloves.
Work in a fume cupboard.
Measure out the indicated quantity of methanoic acid in a measuring cylinder.
CLEAPSS Recipe Book
68 CLEAPSS 2011
60 Nickel sulfate(VI)
Nickel sulfate(VI)-6-water is the most common nickel sulphate salt sold now, but do check the label on the
bottle because the 7-water hydrate is also available.

Formula: NiSO
4
.6H
2
-1

General Hazards The hazards of nickel salts are being constantly reviewed. Sensitisation may be a
problem for some people. See Hazcard 65B.

Procedure for preparing 100 ml of 0.1 M of nickel(II) sulfate(VI) solution
Dissolve 2.63 g of nickel(II) sulfate(VI)-6-water in 70 ml of water.
The solution should be labelled TOXIC.
CLEAPSS Recipe Book
CLEAPSS 2011 69
61 Nitric(V) acid
The commercial solution is about 70% (w/v) and has a specific gravity of 1.42. Its concentration is
15.8 mol dm
-3
.
Concentrated nitric(V) acid should be a colourless solution and stored in dark bottles. When exposed to light,
the solution goes brown and TOXIC nitrogen dioxide gas is produced (see Hazcard 68). Nitrogen dioxide will
leak from bottles and the solution will become less concentrated.
Also available from some suppliers is 95-100% (fuming) nitric(V) acid, which is even more hazardous as it is
unstable. It is unsuitable for schools to store this for long periods. Contact CLEAPSS if you find some.
Do not make dilute solutions for the first time without seeking practical advice from a more-experienced
colleague.

Formula: HNO
3
-1

General Hazards See Hazcard 67. If in contact with the skin, this becomes yellow and layers of skin
peel off.

Concentration
required
label
500 1000 2500
0.01 M Ten-fold dilution of a 0.1 M solution with water -
0.1 M Ten-fold dilution of a 1 M solution with water IRRITANT
0.4 M 13 25 63 IRRITANT
1 M 32 63 158 CORROSIVE
5M 158 317 792 CORROSIVE

Procedure
Wear goggles (a face shield is preferable when handling large volumes) and chemical-resistant
gloves.
Work in a fume cupboard.
Measure out the indicated volume of concentrated nitric(V) acid in a measuring cylinder.
Add the liquid to about two thirds of the final volume of water in suitable beaker or laboratory jug.
Stir the solution (which will become warm).
Pour the solution into a labelled bottle and mix well. Add a hazard warning if appropriate.
CLEAPSS Recipe Book
70 CLEAPSS 2011
62 Nylon rope experiment
Although used in the past, tetrachloromethane (carbon tetrachloride) (see Hazcard 100) and
1,1,1-trichloroethane (see Hazcard 103), should no longer be chosen as the solvent. An alternative is
cyclohexane.
Hexanedioyl chloride (adipoyl chloride) may be used but is more unstable in storage, so it is normally better
to buy decanedioyl (sebacoyl) chloride.
The acid chlorides are liquids at room temperature. They can be weighed out in a suitable container.
Suppliers also provide the prepared solutions. These are used directly. Do check the labels.

General Hazards The acid chlorides and hexane-1,6-diamine are CORROSIVE (see Hazcards 3B & 41)
and cyclohexane is HIGHLY FLAMMABLE & HARMFUL (see Hazcard 45B).

Procedure using cyclohexane as the solvent
Wear eye protection. Wear disposable nitrile gloves when pulling out the nylon thread. The room
should be well ventilated and there must be no sources of ignition.
Dissolve 2.2 g of hexane-1,6-diamine in 50 ml of distilled water in a beaker (label it A).
Dissolve 1.5 g of decanedioyl chloride or hexanedioyl chloride in 50 ml of cyclohexane in another
beaker (label it B) and add HIGHLY FLAMMABLE & HARMFUL warning signs.

Large-scale version
Carefully pour the solution from beaker B onto solution A down a glass rod. The two liquid phases do
not mix; resist any attempt to stir the mixture at all.
Use forceps to pull out the nylon formed at the interface of the two solutions.
Disposal: see Handbook section 7.5.

Small-scale version
Place sufficient solution A to cover the base in a Petri dish.
Carefully pour solution B onto solution A in a Petri dish down a glass rod. The two liquid phases do not
mix; resist any attempt to stir the mixture at all.
Use forceps to pull out the nylon formed at the interface of the two solutions.
The thread can pulled over glass rods or pulleys smeared with a lubricant such as WD40 (see pictures
below.)
Disposal: see Handbook section 7.5.

It is also possible to reduce the scale further and use 5 ml beakers. This is particularly suitable for use by
students.

CLEAPSS Recipe Book
CLEAPSS 2011 71
63 Oscillating reactions
More information about these reactions can be found in Chemical Demonstrations (Vol 2), Shakashiri, 1985,
University of Wisconsin Press, ISBN 0299101304. The entire four volume series is a good source of
information.
The blue-bottle reaction
Wear goggles to prepare the sodium hydroxide solution.
Wear eye protection for the experiment.
In a 500 ml flask, dissolve 10 g of glucose in 300 ml of 0.4 M sodium hydroxide
solution. Add 6 drops of a 0.2% aqueous methylene blue solution. Fit a rubber
bung.
When the solution becomes colourless, it can be shaken to turn it blue again after
which it will again turn colourless.
Other dyes and mixtures of dyes that can be used are:
1% resazurin solution (pale blue to purple pink).
1% indigo carmine (yellow to red brown and to green with vigorous shaking).
0.2% phenosafranine (pink to colourless).
Phenosafranine and methylene blue (pink to purple and to blue with vigorous
shaking) - amounts need to be found by trial and error.
Methyl red and methylene blue (yellow to green) amounts need to be found by
trial and error.
See Hazcards 32 &
91. The solution
should be labelled
IRRITANT.
The Belousov-Zhabotinski (B-Z) reaction
All reagents in the Belousov-Zhabotinski (B-Z) reactions must be free of chloride ions,
which inhibit the reaction.
Place 100 ml of 1 M sulfuric(VI) acid in a beaker on a magnetic stirrer.
Add 2.86 g of malonic acid. Let it dissolve.
Add 1.04 g of potassium bromate(V). Let it dissolve.
Add 0.11 g of cerium(IV) ammonium nitrate(V) (IRRITANT). Let it dissolve.
Add about 0.5 ml of ferroin indicator (see Recipe Sheet 49). If the red/green
oscillations slow down, more potassium bromate(V) can be added.
See Hazcards 6B,
80 & 98A.
The Briggs-Rausher (B-R) reaction
All reagents in the Briggs-Rausher (B-R) reaction must be free of chloride ions, which
inhibit the reaction.
Solution A: 10 vol hydrogen peroxide solution.
Solution B: 6% (w/v) potassium iodate(V) solution in 0.1 M sulfuric(VI) acid.
Solution C: 1% starch solution.
Solution D: a solution containing 2% (w/v) malonic acid and 0.45% (w/v)
manganese sulfate(VI).
Add the solutions to a beaker on an magnetic stirrer in the following order and
quantities: 20 ml of solution A, 20 ml of solution B, 1 ml of solution C and 20 ml of
solution D.
See Hazcards 6B,
50 & 80. The
diluted solutions do
not require a
hazard warning.
D. The nitrogen gas oscillating reaction
Use a fume cupboard (nitrogen dioxide, VERY TOXIC, is produced).
Solution A: Dissolve 2.6 g of ammonium sulfate(VI) in 10 ml of 0.2 M sulfuric(VI)
acid.
Solution B: Dissolve 2.8 g of sodium nitrate(III) (nitrite) in 10 ml of water.
Mix the two solutions in a 100 ml beaker and stir quickly. Transfer to a 50 ml
measuring cylinder and after a minute the oscillations become quite distinct.
See Hazcards 68 &
93.

CLEAPSS Recipe Book
72 CLEAPSS 2011
64 Oxygen gas
Oxygen is prepared by adding hydrogen peroxide solution onto a catalyst, manganese(IV) oxide.
The reaction is very vigorous and exothermic and chemicals have been known to shoot up the thistle funnel
or the bung has been forced out of the flask. The rate of the reaction depends on the surface area of the
catalyst. It is better to use granules rather than a fine powder. If only powder is available, then only a small
amount is required.
Potassium iodide (1 g) produces oxygen at a steadier rate. The solution first saturates itself with oxygen and
then the gas is released.
Hydrogen peroxide decomposes slowly during storage. Suppliers add an inhibitor to slow the process, but
once this is used up, decomposition accelerates. If more hydrogen peroxide than expected is required, then
this is a sure sign that the hydrogen peroxide is old and new stock is required.
5 ml of 100 vol solution of hydrogen peroxide produces 500 ml of oxygen, ie, 5 x 100 ml.

General Hazards See Hazcards 50, 60 & 69.

In the 250 ml Bchner flask, add 0.1 g of manganese(IV) oxide (HARMFUL) powder or 1 g of granules.
Fill the measuring cylinder with water ready to collect the first 250 ml of gas produced. The measuring
cylinder allows you to be confident that apparatus is well flushed out before oxygen is collected.
Pour at least 25 ml of cold water, enough to cover the bottom, into the thistle funnel.
Now add 5 ml of 100 vol hydrogen peroxide (HARMFUL).
Once the measuring cylinder is full of gas, remove it and replace with a gas jar full of water.
Keep collecting gas jars of gas. When full of gas, place a glass cover on the gas jar.
If the rate of gas being produced slows down, add further 5 ml portions of 100 vol hydrogen peroxide.

Add 5 ml of 100 vol
hydrogen peroxide
250 ml measuring
cylinder
250-ml
Bchner
Flask
CLEAPSS Recipe Book
CLEAPSS 2011 73
65 Phosphoric(V) acid
Also known as orthophosphoric acid.
The usual concentrated commercial solution is about 85% (w/v).
-3
.

Formula: H
3
PO
4
Density: 1.7 g cm
-3
Molar mass: 98 g mol
-1

General Hazards See Hazcard 72.

Concentration
required
label
500 1000 2500
0.1 M 3.5 7.0 17.0 -
0.5 M 17 34 85 -
1 M 34 68 170 IRRITANT

Procedure
Wear goggles and chemical-resistant gloves.
Measure out the indicated volume of concentrated phosphoric(V) acid in a measuring cylinder.
Add the liquid to about two thirds of the final volume of water in a suitable beaker or laboratory jug.
Stir the solution.

CLEAPSS Recipe Book
74 CLEAPSS 2011
66 Plant mineral requirement solutions
A Sachs culture solution (complete recipe)
Dissolve the following salts in 1000 ml of pure water:
0.25 g of calcium sulfate(VI)-2-water
0.25 g of calcium phosphate(V)-2-water [CaH
4
(PO
4
)
2
.2H
2
O]
0.25 g of magnesium sulfate(VI)-7-water
0.08 g of sodium chloride
0.70 g of potassium nitrate(V)
0.005 g of iron(III) chloride-6-water

B Sachs culture solution with mineral deficiencies
Deficient in calcium: 0.2 g of potassium sulfate(VI) replaces calcium sulfate(VI)-2-water and
0.71 g of sodium dihydrogenphosphate(V)-2-water replaces calcium
phosphate(V) in recipe A.
Deficient in iron: Omit iron(III) chloride-6-water in recipe A.
Deficient in magnesium: 0.17 g of potassium sulfate(VI) replaces magnesium sulfate(VI) in recipe A.
Deficient in nitrogen: 0.52 g of potassium chloride replaces potassium nitrate(V) in recipe A.
Deficient in phosphorus: 0.16 g of calcium nitrate(V)-4-water replaces calcium phosphate(V) in
recipe A.
Deficient in potassium: 0.59 g of sodium nitrate(V) replaces potassium nitrate(V) in recipe A.
Deficient in sulfur: 0.16 g of calcium chloride replaces calcium sulfate(VI) and 0.21 g of
magnesium chloride-6-water replaces magnesium sulfate(VI) in recipe A.

C Knops culture solution
Solution 1
Dissolve the following salts in 1000 ml of pure water:
1 g of magnesium sulfate(VI)
1 g of potassium nitrate(V)
1 g of dipotassium hydrogenphosphate(V) [K
2
HPO
4
]

Solution 2
Dissolve 4 g of calcium nitrate(V) in 1000 ml of pure water.

Add solution 1 to solution 2 before use. This recipe is suitable for algae. For flowering plants, further dilution
by 2, 3 or 4 times may produce better results. This should be determined by experiment.
CLEAPSS Recipe Book
CLEAPSS 2011 75
67 Potassium and sodium phosphates
These solutions are used to make buffer solutions (see Recipe Sheet 18 for Buffers). Masses are given to 3
decimal places but for many purposes, such accuracy is not required. Adjust to 2 decimal places.
Make sure you are using the correct salt. It is best to check with the formula (see below).
Sodium and potassium dihydrogen phosphate(V) solutions are best prepared just before use. During storage
there may be mould growth.

Potassium dihydrogen
phosphate(V)
KH
2
PO
4
0.2 M Dissolve 27.218 g of the solid in 500 ml of water
and add water to 1000 ml.
Sodium dihydrogen
phosphate(V)
NaH
2
PO4

Sodium dihydrogen
phosphate(V)-1-water
NaH
2
PO
4
.H
2
O 0.2 M Dissolve 27.598 g of the solid in 500 ml of water

Dipotassium hydrogen
phosphate(V)
K
2
HPO
4
K
2
HPO
4
.3H
2
Disodium hydrogen
phosphate(V)
Na
2
HPO
4
Disodium hydrogen
Na
2
HPO
4
.7H
2
0 0.2 M Dissolve 53.614 g of the solid in 500 ml of water

Potassium
phosphate(V)
K
3
PO
4
Sodium phosphate(V) Na
3
PO
4
Sodium phosphate(V)-
12-water
Na
3
PO
4
.12H
2

CLEAPSS Recipe Book
76 CLEAPSS 2011
68 Potassium chloride
Formula: KCl Molar mass: 74.55 g mol
-1

Concentration
required
label
100 250 1000
1.0 M 7.46 18.64 74.55 -
3.0 M (used for
filing pH probes)
22.34 55.91 223.65 -
Saturated (20 C) 40 100 400 -

Procedure
Measure out the indicated quantity of potassium chloride.
CLEAPSS Recipe Book
CLEAPSS 2011 77
69 Potassium chromate(VI)
Formula: K
2
CrO
4
-1


Concentration
required
label
100 250 1000
0.01 M Ten-fold dilution of the 0.1 M solution TOXIC
0.1 M 1.94 4.86 19.42 TOXIC
1.0 M 19.42 48.58 194.19 VERY TOXIC
Saturated (20 C) 68 158 630 VERY TOXIC

Procedure
Wear chemical-resistant gloves and goggles. Use a fume cupboard if the solid is a fine powder.
Measure out the indicated quantity of potassium chromate(VI).
Pour into a labelled bottle. Add the hazard warning.

CLEAPSS Recipe Book
78 CLEAPSS 2011
70 Potassium dichromate(VI)
Potassium dichromate solutions are best made up in 0.1 M sulfuric(VI) acid rather than in pure water.
This can be used as acidified potassium dichromate(VI) solution.
The solution is used to test for the presence of alcohols. See Recipe Sheet 102 for Test reagents for organic
functional groups.

Formula: K
2
Cr
2
O
7
-1


Concentration
required
label
100 250 1000
0.01 M Ten-fold dilution of the 0.1 M solution with 0.1 M sulfuric(VI) acid TOXIC
0.1 M 2.94 7.36 29.42 TOXIC
Saturated (20 C) 12 30 120 VERY TOXIC

Procedure
Measure out the indicated quantity of potassium dichromate(VI).
Add the solid to about two thirds of the final volume of 0.1 M sulfuric(VI) acid in a beaker or laboratory
jug.
0.1 M sulfuric(VI) acid to the required level.
Pour into a labelled bottle. Add the hazard warning.
CLEAPSS Recipe Book
CLEAPSS 2011 79
71 Potassium hydroxide
Solutions absorb carbon dioxide on standing. Solutions lower than 0.1 M are most significantly affected and
should, therefore, be freshly made.
Potassium hydroxide solution is best stored in plastic screw-cap bottles. It is known to etch glass when
stored for long periods and polystop caps can allow carbon dioxide to enter.
Equipment with glass joints, eg, burettes and bottles, can seize up over time.

Formula: KOH Molar mass: 56.11 g mol
-1

General Hazards See Hazcard 91. When added to water so much heat is evolved that boiling could
occur.
Do not make this solution for the first time without seeking practical advice from a
more experienced colleague. A choking mist is often formed as the solid dissolves in
water. While this is not a serious safety risk, it is unpleasant and it is wise to use a
fume cupboard where possible. If saturated potassium hydroxide has to be prepared
after carrying out an exhaustive risk assessment, then start from 5 M potassium
hydroxide solution and add pellets a little at a time. Do not store this solution.

Concentration
required
label
100 250 1000
0.1 M Ten-fold dilution of the 1 M solution IRRITANT
0.4 M 2.24 g 5.61 g 22.44 g IRRITANT
1.0 M 5.61 g 14.03 g 56.11 g CORROSIVE
5.0 M 28.06 g 70.14 g 280.55 g CORROSIVE

Procedure
Measure out the indicated quantity of potassium hydroxide.
Add the solid in stages to about two thirds of the final volume of water in a beaker or laboratory jug. If
concentrated solutions are being made, ice should be used in place of water.
Stir carefully to dissolve before adding the next group of pellets. It may be necessary to cool the
solution between additions. (Ice could be added.)
Pour into a labelled plastic bottle. Add a hazard warning if appropriate.

A 10% (w/v) alcoholic solution of potassium hydroxide

Measure out 10 g of potassium hydroxide.
Place 70 ml of the alcohol (ethanol or methanol) in a glass 250 ml beaker.
Insert a magnetic stirrer bar and place the beaker on a magnetic stirrer.
Add the potassium hydroxide solution in 3 parts, allowing the solid to dissolving before adding the next
amount. The process is exothermic but not as extreme as that with water.
Place the solution in a 100 ml measuring cylinder and make up to the 100 ml mark with the alcohol.
Pour into a labelled plastic bottle. Label the solution CORROSIVE and HIGHLY FLAMMABLE.

CLEAPSS Recipe Book
80 CLEAPSS 2011
72 Potassium iodide
Solutions of potassium iodide often go yellow on keeping. If this occurs, add 0.1 M sodium thiosulfate
solution dropwise, or a crystal of the solid with stirring, until the solution becomes colourless.

Formula: KI Molar mass: 166.00 g mol
-1

Concentration
required
label
100 250 1000
0.1 M 1.66 4.15 16.60 -
1.0 M 16.60 41.50 166.00 -
Saturated (20 C) 150 375 1500 -

Procedure
Measure out the indicated quantity of potassium iodide.

CLEAPSS Recipe Book
CLEAPSS 2011 81
73 Potassium manganate(VII)
Also known as potassium permanganate.
Solutions must be acidic. Solutions are safer made up in dilute sulfuric(VI) acid rather than adding
concentrated sulfuric(VI) acid at a later stage.
Solutions do not keep well unless the container is scrupulously clean. They slowly reacts with water, forming
manganese(IV) oxide which badly stains glass and plastic equipment.
Solutions are best kept in dark bottles, shielded from light. Light increases the rate of decomposition and
staining of equipment.
Equipment can be cleaned by filling the bottle either with 1 M sulfuric(VI) acid with a small amount of
hydrogen peroxide solution (20 or 100 vol) or, if this does not work, add 2 M hydrochloric acid and leave it for
several hours (or days) in a fume cupboard.
Use a 0.002 M solution for carrying out tests for unsaturation in alkenes.
It is not possible to make a 1 M solution.

Formula: KmnO
4
-1


Concentration
required
Conc of acid
for dilution
warning label
500 1000 2500
0.002 M 0.1 M Ten-fold dilution of the 0.02 M solution
with 0.1 M sulphuric(VI) acid
-
0.02 M 0.1 M 1.58 3.16 7.90 -
0.1 M 1 M 7.90 15.80 39.51 IRRITANT
Saturated 1 M 32 64 160 IRRITANT

Procedure
It would be advisable to wear gloves as the chemical stains the skin brown.
Measure out the indicated quantity of potassium manganate(VII).
Add the solid to about two thirds of the final volume of the suggested dilute sulfuric(VI) acid in a
beaker or laboratory jug.
Stir to dissolve. It is better not to heat but dissolving can take some time, and it is difficult to see when
the solid has all dissolved.
dilute sulfuric(VI) acid to the required level.
CLEAPSS Recipe Book
82 CLEAPSS 2011
74 Preservatives used after fixing biological specimens
Preservatives prevent chemical decomposition of the fixed material (see Recipe Sheet 41).
More information about the procedures for fixing and preserving specimens can be found in the CLEAPSS
Handbook, section 15.8.

Preserving animal
tissue (Recipe 1)
Add 10 ml of odourless preservative
(eg, Phenoxetol or Opresol) to 50 ml
of propane-1,2-diol. Dilute this
mixture to 100 ml with purified water.
(Another version of this recipe
includes and additional 1 g of sodium
chloride.)
Wear eye protection. See
Hazcard 37.
Animal tissue should not be
preserved for more than 1 month in
these solutions unless it is fixed
first. Label the solution IRRITANT.
Preserving animal
tissue (Recipe 2)
Mix together 10 g of 4-methoxybenz-
aldehyde (p-anisaldehyde), 50 ml of
Teepol and 50 ml of propane-1,2-diol
and make up this mixture to 1000 ml
with purified water.
Wear eye protection. See
Hazcard 37.
Animal tissue should not be
preserved for more than 1 month in
these solutions unless it is fixed
first. Label the solution IRRITANT.
4-Methoxybenzaldehyde is
HARMFUL if swallowed. Label the
solution HARMFUL.
Preserving plant tissue Use the formalin-aceto-alcohol (FAA)
recipe on the fixative recipe sheet 41.
To preserve the red or green colour
add copper(II) sulfate(VI)-5-water
crystals; the amount is not too
critical.
Use a fume cupboard. Wear eye
protection.
For ethanol; see Hazcard 40A.
For methanal, see Hazcard 63.
For ethanoic acid, see Hazcard 38.
For copper(II) sulfate(VI), see
Hazcard 27C. Label the solution
HIGHLY FLAMMABLE, IRRITANT and
HARMFUL.
Kaiserlings
preservative
Add 36 g of sodium ethanoate, 5 ml
of a 5% camphor solution in ethanol
and 72 ml of propane-1,2,3-triol
(glycerol). Add this mixture to 120 ml
of purified water.
Camphor is HARMFUL. For propane-
1,2,3-triol, see Hazcard 37. For
sodium ethanoate see Hazcard
38A. for ethanol, see Hazcard 40A.
Label the solution HARMFUL and
HIGHLY FLAMMABLE.

CLEAPSS Recipe Book
CLEAPSS 2011 83
75 Ringers and other saline solutions for physiological use
Saline solutions are suitable for temporary microscopial preparations and short experiments.
For more prolonged investigations, a suitable Ringers solution should be used.

Ringers solutions
For frogs Dissolve 6.50 g of sodium chloride, 0.12 g of calcium chloride-6-water (IRRITANT), 0.14 g of
potassium chloride and 0.20 g of sodium hydrogencarbonate in 1000 ml of water.
For mammals Dissolve 8 g of sodium chloride, 0.2 g of calcium chloride-6-water (IRRITANT), 0.2 g of
potassium chloride, 0.19 g of magnesium chloride-6-water, 0.05 g of sodium
dihydrogenphosphate and 1 g of sodium hydrogencarbonate in 1000 ml of water.
For locusts
and other
insects
Dissolve 7.60 g of sodium chloride, 0.22 g of calcium chloride-6-water, 0.75 g of potassium
chloride, 0.19 g of magnesium chloride-6-water, 0.48 g of sodium dihydrogen phosphate,
and 0.37 g of sodium hydrogencarbonate in 1000 ml of water.
For
earthworms
Dissolve 6 g of sodium chloride, 0.2 g of calcium chloride-6-water, 0.12 g of potassium
chloride and 0.1 g of sodium hydrogencarbonate in 1000 ml of water.
For marine
crustaceans
Dissolve 31 g of sodium chloride, 1.37 g of calcium chloride-6-water (IRRITANT), 0.99 g of
potassium chloride, 2.35 g of magnesium chloride-6-water, 0.48 g of sodium
dihydrogenphosphate, and 0.22 g of sodium hydrogencarbonate in 1000 ml of water.

Saline solutions
For invertebrate tissues Dissolve 7.5 g of sodium chloride in 1000 ml of water.
For amphibian tissues Dissolve 6.4 g of sodium chloride in 1000 ml of water.
For mammalian tissues Dissolve 9 g of sodium chloride in 1000 ml of water.
For mammalian blood Dissolve 6 g of sodium chloride in 1000 ml of water.

CLEAPSS Recipe Book
84 CLEAPSS 2011
76 Sandells solution
Sandells solution is a safer alternative than Fehlings testing for sugars.
Sandells solution (which is little known) can be stored for a long time.
The advantage in its use is that the concentration of sodium hydroxide is 0.4 M in the solution, much lower
than in Fehlings solution B, which is close to 4M.
The method, by A. Sandell, was published in the Journal of Chemical Education, April 1994, p346. SSERC
has shown that this solution is stable enough to store and works with both aldehydes and monosaccharides.
It is used in the same way as Benedicts solution.

General Hazards Copper(II) sulfate(VI) crystals are HARMFUL if swallowed (see Hazcard 27C).
2M sulfuric(VI) acid, solid sodium hydroxide and 0.5 M sodium hydroxide solution
are CORROSIVE (see Hazcards 91 and 98A). EDTA (disodium salt) is low hazard
(see Hazcard 3B).

Procedure to make 100 ml of Sandells solution
Wear goggles when preparing the solution.
Measure out 0.80 g of copper(II) sulfate(VI)-5-water and 1.80 g of EDTA (disodium salt), and dissolve
in 80 ml of water.
Add 20 ml of 2 M sodium hydroxide (CORROSIVE).
Make up to 100 ml with water. Label the bottle IRRITANT.
CLEAPSS Recipe Book
CLEAPSS 2011 85
77 Silver nitrate(V)
This is probably the most expensive chemical used in schools; use it with care, retrieving unused stock.
Pure water (ie, distilled or deionised) should be used to avoid cloudiness of solutions.
Tollens reagent [ammoniacal silver nitrate(V) solution], (see Recipe sheet 102) should not be stored. It
should be made and disposed of in situ by the students.

Formula: AgNO
3
-1

General Hazards See Hazcard 87. The solid and more-concentrated solutions will stain skin and other
organic material black. This stain is difficult to remove. Work surfaces also stain.

Concentration
required
label
100 250 1000
0.01 M Ten-fold dilution of the 0.1 M with water -
0.1 1.70 4.25 16.99 -
0.2 3.40 8.50 33.98 IRRITANT
0.5 8.49 21.23 84.94 CORROSIVE
Saturated (20 C) 220 550 2200 CORROSIVE

Procedure
Measure out the indicated quantity of silver nitrate(V).
CLEAPSS Recipe Book
86 CLEAPSS 2011
78 Slime
Different sources of polyvinyl alcohol vary in their average molar mass; some are about 115000 g mol
-1
,
while others are much less, at around 17000 g mol
-1
(and less expensive). The larger the average molar
mass, the better the slime.
PVA wood glue is cheaper still but contains polyvinyl acetate and other chemicals such as binders. This will
affect the nature of the slime and it may become more solid.
Custard powder can also be used with PVA glue to form bouncing custard balls.
Slime is a gel. It is thought that borate ions react with hydroxyl groups in the polymer of vinyl alcohol to form
cross links, with the elimination of water. These cross links probably involve hydrogen bonds which
continually form and break under flow.
The sliminess of the gel can be adjusted by altering the amount of sodium borate used. Viscosity changes
can be detected by timing the passage of a ball bearing through the slime in a measuring cylinder.
If acid is added to the slime, the gel collapses to give a free-flowing solution.
Although the slime can be stored in a labelled sealed bag and placed in a refrigerator, it can develop mould if
kept for long periods.
Disposal: Add enough 1 M sulfuric(VI) acid to just allow the solution to become free-flowing, dilute and pour
down a foul-water drain.
These chemicals, and the made-up slime, should not be taken home.

General Hazards See Hazcard 14. However, since publication of Hazcards, the toxicity of boron
compounds has been the subject of a considerable review and debate in the
European Union. Some suppliers will now label the bottle TOXIC with R63 (Possible
risk of harm to the unborn child) warning. However only solutions above 8% (w/v)
will be labelled TOXIC.
Sodium tetraborate should be weighed in a fume cupboard to avoid raising dust.

Procedure using polyvinyl alcohol (PVA)

PVA with a molar mass above 85,000 g mol
-1
PVA with a molar mass below 85,000 g mol
-1

Wear eye protection. Hot water from a kettle may be useful to start with. Those with sensitive skin may be
affected by the weak alkalinity of the solutions. In this case, disposable gloves should be worn.
Pour about 100 ml of water (no hotter than 90 C)
into a 400 ml beaker, add 4 g of high-mass
polyvinyl alcohol, stirring rapidly. Add food
colouring and/or fluorescent dye, eg, fluorescein,
for added (disgusting) effect.
Heat to 90 C (but do not boil) and keep stirring
as required until the polymer dissolves. Allow the
solution to cool. All of this may take some time
(Solution A.).
Dissolve 0.80 g of sodium tetraborate-10-water
(0.42 g of anhydrous sodium tetraborate) in
20 ml of water. (Solution B.)
Pour about 100 ml of water (no hotter than 90 C)
into a 400 ml beaker, add 8 g of low-mass
polyvinyl alcohol, stirring rapidly. Add food
colouring and/or fluorescent dye, eg, fluorescein,
for added (disgusting) effect.
Heat to 90 C (but do not boil) and keep stirring as
required until the polymer dissolves. Allow the
solution to cool. All of this may take some time
(Solution A.).
Dissolve 1.6 g of sodium tetraborate-10-water
(0.84 g of anhydrous sodium tetraborate) in
20 ml of water. (Solution B.)
Add solution B to A with vigorous stirring. Let the gel form before removing it and washing with water.

Using PVA glue:
Dilute it to about 25%; there is no need to heat it but mix it well with water. Add sodium tetraborate as
prepared above for use with low-mass PVA. Change the concentration of PVA glue to alter the slime
quality.
CLEAPSS Recipe Book
CLEAPSS 2011 87
79 Soap and bubble solutions
Alcoholic soap solution can be titrated against tap water in a stoppered bottle until a froth, stable for 15
seconds, is obtained. The concentration of soap solution and/or volume of water involved in the titration will
depend on the hardness of the local tap water. Solutions should be tested before being presented to a class
so that minor adjustments to the formulation can be made.
The alcoholic soap solution is called either Clarkes or Wanklyns soap solution.

General Hazards See Hazcards 37 & 40A. Do not heat soap solutions made up in ethanol with a
naked flame. Use a hot plate or surround the beaker with hot water.

Procedure to make a soap solution suitable for titrating
Dissolve 5 g of soap flakes (eg, Lux) in 500 ml of ethanol by stirring the suspension on a hot plate.
This may take some time.
After cooling, pour the solution into a 1 litre volumetric flask and dilute to 1000 ml with pure water. Mix
the solution well.
The titration with tap water to obtain a permanent froth should be rehearsed so that adjustments to the
volume of tap water or concentration of the soap solution can be made.

Bubble mixture
Recipes may differ with different makes of liquid detergent.
The recipe below works best using Fairy Liquid. (other detergents will also work but you may need to
experiment with quantities.

Procedure to make a bubble solution
Mix together by volume, 10 parts liquid detergent, 85 parts pure water and 5 parts propane-1,2,3-triol
(glycerol).
CLEAPSS Recipe Book
88 CLEAPSS 2011
80 Sodium carbonate
Sodium carbonate is sold both as anhydrous sodium carbonate or sodium carbonate-10-hydrate.
Anhydrous sodium carbonate absorbs water from the atmosphere (deliquescence).
Hydrated sodium carbonate loses water to the atmosphere (efflorescence); look for a white powder as
opposed to a glassy solid.
Do not use technical grade sodium carbonate to make up solutions. There is a lot of sediment.
Solutions may be cloudy if hard tap water is used.

Formula: Na
2
CO
3
-1
Formula: Na
2
CO
3
.10H
2
-1


Anhydrous sodium carbonate
Concentration
required
label
250 500 1000
0.05 M Two-fold dilution of 0.1 M solution -
0.1 M 2.65 5.30 10.60 -
0.5 M 13.25 26.50 53.00 -
1 M 26.50 53.00 105.99 -
Saturated 55 110 220 IRRITANT

Hydrated sodium carbonate
Concentration
required
label
250 500 1000
0.05 M Two-fold dilution of 0.1 M solution -
0.1 M 7.15 14.31 28.61 -
0.5 M 35.77 71.54 143.07 -
1 M 71.54 143.07 286.14 -
Saturated 125 250 500 IRRITANT

Procedure
Measure out the indicated quantity of anhydrous or hydrated sodium carbonate.
CLEAPSS Recipe Book
CLEAPSS 2011 89
81 Sodium chlorate(I) solution
Also known as sodium hypochlorite solution, sodium chlorate(I) or bleach.
Do not get confused with the solid sodium chlorate(V).
The solution sold by laboratory suppliers and described as 10-14% available chlorine contains 100 000 to
140 000 ppm of chlorine and has a molarity of 1.5 M.
It has a limited shelf-life and may not be as active after a year in storage. Some technicians use domestic
products such as bleach but please read the containers carefully as some bleaches use hydrogen peroxide
and not sodium chlorate(I). Domestic bleach is usually 0.5 M or less and represents poorer value per mole
but may be suitable if only needed infrequently. However, there are also thickeners and detergents in
domestic bleach that may cause problems. See also Handbook section 15.12. Solutions used for disinfecting
should always be freshly prepared.
If using this as a disinfectant, purchase or prepare fresh stocks after 3 months or so.

Formula: NaClO Molar mass: 74.45 g mol
-1


Concentration required Preparation Use
1% 10000 ppm 0.15 M See below for procedure.
No need for further dilution.
Disinfectant for blood spills or
dirty conditions.
0.25% 2500 ppm 0.038 M Mix, by volume, 1 part of the 1%
solution with 3 parts of water.
Disinfectant for micro-biological
discard pots.
0.1% 1000 ppm 0.015 M Mix by volume, 1 part of the 1%
solution with 9 parts of water.
Disinfectant for general use, eg,
sterilising solution.
0.01% 100 ppm 0.0015 M Dilute the 0.1% solution further
by mixing by volume, 1 part of
the 0.1% solution with 9 parts of
water.
Disinfectant for sterilising
mouthpieces, swabbing skin.

Procedure to make 1000 ml of 1% available chlorine solution
Wear gloves and goggles or a face shield.
Measure out 100 ml of 10-14% sodium chlorate(I) solution into a 1000 ml measuring cylinder.
Add water to the 1000 ml level.
Pour the solution into an appropriate labelled bottle and mix well. The solution is low hazard.
Depending on use, dilute, if necessary, from a 1% solution according to the table above.
CLEAPSS Recipe Book
90 CLEAPSS 2011
82 Sodium chloride
Formula: NaCl Molar mass: 58.44 g mol
-1

Concentration
required
label
100 250 1000
0.5 M 2.92 7.31 29.22 -
1.0 M 5.84 14.61 58.44 -
Saturated (20 C) 37 93 370 -

Procedure
Measure out the indicated quantity of sodium chloride.
CLEAPSS Recipe Book
CLEAPSS 2011 91
83 Sodium ethanoate
Sodium ethanoate is sold as the anhydrous salt or as the trihydrate.

Anhydrous sodium ethanoate
Formula: NaCH
3
COO Molar mass: 82.03 g mol
-1

Concentration
required
label
100 250 1000
0.2 M 5-fold dilution of the 1 M solution -
1.0 M 8.20 20.51 82.03 -
Saturated (20 C) 50 125 500 -

Hydrated sodium ethanoate
Formula: NaCH
3
COO.3H
2
-1

Concentration
required
label
100 250 1000
0.2 M 5-fold dilution of the 1 M solution -
1.0 M 13.61 34.02 136.08 -
Saturated (20 C) 80 200 800 -

Procedure
Measure out the indicated quantity of sodium ethanoate.
CLEAPSS Recipe Book
92 CLEAPSS 2011
84 Sodium hydrogencarbonate
Formula: NaHCO
3
-1

Concentration
required
label
100 250 1000
0.5 4.20 11.50 42.00 -
1.0 M 8.40 21.00 84.01 -
Saturated (20 C) 12 30 120 -

Procedure
Measure out the indicated quantity of sodium hydrogencarbonate.
Stir to dissolve. Do not warm as decomposition may take place.

Sodium hydrogencarbonate - alkaline pyrogallol solution for absorbing or removing
oxygen gas
Although safer than using sodium hydroxide with pyrogallol (benzene-1,2,3-triol), the absorption of oxygen is
slower and may be too slow for some procedures In such circumstances, there is no alternative to the use of
sodium hydroxide. The procedure described below limits the absorption of oxygen from the air.


Procedure
Wear eye protection when preparing the solution.
Prepare a saturated solution of sodium hydrogencarbonate using freshly-boiled pure water.
Place a wide-bore glass tube into the water and then pour liquid paraffin onto the waters surface.
Add a few crystals of benzene-1,2,3-triol (pyrogallol) down the glass tube and then slowly withdraw it.
(1 g of benzene-1,2,3-triol is capable of absorbing 190 cm
3
of oxygen.)

CLEAPSS Recipe Book
CLEAPSS 2011 93
85 Sodium hydroxide
Sodium hydroxide solution is better stored in plastic screw-cap bottles. It is know to etch glass when stored
for long periods and polystop caps can allow carbon dioxide to enter.
Equipment with glass joints, eg, burettes and bottles, can seize up over time.

Formula: NaOH Molar mass: 40.00 g mol
-1

General Hazards See Hazcard 91. When added to water, heat is evolved such that boiling could
occur. Do not make this solution for the first time without seeking practical advice
from a more-experienced colleague. A choking mist is often formed as the solid
dissolves in water. While this is not a serious safety risk, it is unpleasant and it is
wise to use a fume cupboard where possible. If after carrying out an exhaustive risk
assessment, there is no alternative to preparing saturated sodium hydroxide, then
start from 5 M sodium hydroxide solution and add pellets a little at a time. Do not
store this solution.

Concentration
required
label
250 1000 2500
0.1 M Ten-fold dilution of the 1 M solution IRRITANT
0.4 M 4.00 16.00 40.00 IRRITANT
1.0 M 10.00 40.00 100.00 CORROSIVE
5.0 M 50.00 200.00 500.00 CORROSIVE

Procedure
Measure out the indicated quantity of sodium hydroxide.
Add the solid in stages to about two thirds of the final volume of water in a beaker or laboratory jug. If
concentrated solutions are being made, ice should be used in place of water.
Stir carefully to dissolve before adding the next group of pellets. It may be necessary to cool the
solution between additions. (Ice could be added.)

CLEAPSS Recipe Book
94 CLEAPSS 2011
86 Sodium silicate, the crystal (chemical) garden and silicate gels
The solution is also known as water glass.
It is more convenient to buy a solution which has a specific gravity of about 1.5, rather than the solid, and
dilute it down.
It is, however, possible to make a solution from the solid.
Expensive glassware can be very difficult to clean after using this reagent. In this case, it is better to use old
jam or coffee jars and dispose of them after use.

General Hazards See Hazcard 95B. The solution is strongly alkaline. Consult the relevant Hazcards
when using metal salts to add to the chemical garden.

Procedure to make 250 ml of sodium silicate solution for a chemical garden from the
commercially-available solution
Add 200 ml of commercial sodium silicate to 600 ml of pure water in a 1-litre beaker.
Pour into a labelled bottle. Although the solution is deemed low hazard, it might be prudent to label the
bottle IRRITANT as the solution is strongly alkaline.

Procedure to make 250 ml of 8% sodium silicate solution from the solid
Add 20 g of sodium silicate to a 600 ml beaker and add 250 ml of pure water.
Add a magnetic stirrer bar and place the mixture on a heater/stirrer.
Maintain a temperature of about 45 C until the solid appears to dissolve.
Filter through Whatman No.1 filter paper.
Pour into a labelled bottle. Although the solution is deemed low hazard, it might be prudent to label the
bottle IRRITANT as the solution is strongly alkaline.

The crystal (chemical) garden
Wear eye protection and consider wearing disposable nitrile gloves.
Pour the prepared sodium silicate into a 600 ml beaker (a labelled coffee jar is also suitable), cover it
with a clock glass or sheet of glass (check edges are not sharp) and allow the liquid to settle. (After
adding crystals, it is better not to move the container so any moving should be done at this stage.)
Now add small (eg, rice-grain size) crystals of manganese(II) sulfate(VI), copper(II) sulfate(VI),
chromium(III) chloride, iron(II) sulfate(VI), iron(III) chloride, cobalt(II) chloride, tin(II) chloride,
aluminium sulfate(VI), nickel(II) sulfate(VI) or any other soluble salt you wish to try.
Leave the container undisturbed for several days. Label the container IRRITANT.

Growing lead(II) iodide crystals in gels
Dilute 16 ml of commercial sodium silicate solution to 100 ml with pure water (solution A).
Place 15 ml of 1 M ethanoic acid (see Recipe Sheet 39) in a beaker and add 0.67 g of potassium
iodide (solution B).
Add 15 ml of solution A to solution B, stirring vigorously. Pour the mixture into a boiling tube and allow
it to set (5 to 60 minutes).
Dissolve 0.66 g of lead(II) nitrate(V) in 2 ml of water and pour this onto the top of the gel. Label the
container IRRITANT.
Observe the spread of crystals of lead(II) iodide over the next few weeks.
CLEAPSS Recipe Book
CLEAPSS 2011 95
87 Sodium thiosulfate
Also known as hypo and it is usually supplied as hydrated crystals.
Distilled water is naturally acidic (dissolved carbon dioxide) and this may result in the slow formation of sulfur
over several hours. This is avoided by adding a pinch of sodium hydrogencarbonate or sodium sulfate(IV) to
the solution to neutralise the acidity.
A 25% solution is suitable to keep with bromine for use in case of accidents. (See Hazcard 95C.)

Formula: Na
2
S
2
O
3
.5H
2
0 Molar mass: 248.17 g mol
-1

Concentration
required
label
100 250 1000
0.1 M 2.48 6.20 24.82 -
0.5 M 12.41 31.02 124.08 -
1.0 M 24.82 62.04 248.17 -
Saturated (20 C) 75 188 750 -

Procedure
Measure out the indicated quantity of sodium thiosulfate-5-water.
Stir to dissolve. Do not warm the solution.
Add about 1 g of sodium hydrogencarbonate or sodium sulfate(IV) (sulfite) (HARMFUL) for every
1000 ml of solution, to help to preserve it.
Pour the solution into a labelled bottle and mix well.

A saturated solution
The saturated solution requires little water. Weigh out the sodium thiosulfate as indicated and add it to hot
(almost boiling) water with a volume in millilitres equal to about half the weight of solid in grams. Stir to
dissolve and leave to cool to room temperature. If there is no solid present in the cooled solution, add one
crystal of solid to it. If this causes further solid to form, then the solution is saturated and may be left in
contact with the solid. If not, add further solid.

CLEAPSS Recipe Book
96 CLEAPSS 2011
88 Stains for bacterial activity
Gram Stain
For details on
using this stain
see Guidance
Leaflet GL 95
To prepare the crystal violet solution, dissolve 2 g of
the dye in 100 ml of industrial denatured alcohol
(Solution A) and 1 g of ammonium ethanedioate in
100 ml of distilled/deionised water (Solution B).
Add 25 ml of solution A to 100 ml of solution B.
To prepare the Safranin counterstain, dissolve 0.5 g of
safranin in 100 ml of distilled/deionised water.
To prepare Grams (Lugols) iodine, dissolve 1 g of
iodine and 2 g of potassium iodide in 300 ml of distilled
or deionised water.
See Hazcards 32, 36A, 40A
and 85.
IDA is HIGHLY FLAMMABLE.
There should be no sources of
ignition in the vicinity
Wear eye protection during the
preparation. Ensure the room
is well ventilated. Wear
chemical-resistant gloves
when preparing the crystal
violet stain.

CLEAPSS Recipe Book
CLEAPSS 2011 97
89 Stains for cells

Aceto carmine Add 0.5 g of carmine to 55 ml of purified water in a
conical flask. Boil and add 45 ml of concentrated
ethanoic acid. Plug the flask with cotton wool, boil
again, cool and filter.
For ethanoic acid, see
Hazcard 38. Label the stain
CORROSIVE. Wear goggles.
Use a fume cupboard.
Ehrlichs
haematoxylin
for animal
histology
Dissolve 0.64 g of haematoxylin in 32 ml of ethanol
heated on a water bath, and filter. Separately, dissolve
0.32 g of aluminium potassium sulfate(VI) in 32 ml of
purified water. Mix this solution with the haematoxylin
solution and add 32 ml of propane-1,2,3-triol (glycerol)
and 4 ml of glacial ethanoic acid. Leave exposed to
daylight for 6 weeks to ripen or add 1 ml of 0.2 M
potassium manganate(VII) solution.
For aluminium potassium
sulfate(VI), see Hazcard 3B.
For ethanol; see Hazcard 40A.
Hazcard 38. For propane-
1,2,3-triol, see Hazcard 37. For
For potassium manganate(VII),
see Hazcard 81.
Wear eye protection and
disposable nitrile gloves when
making up the solution.
Label the stain HIGHLY
FLAMMABLE.
Eosin for
cytoplasmic
staining
Dissolve 1 g eosin in 100 ml tap water. Used as a
counterstain to Ehrlichs haematoxylin. For alcoholic
solution, substitute 75% ethanol for water.
Wear eye protection. Low
hazard unless ethanol is used
as the solvent.
Feulgen's stain
for DNA
Dissolve 0.45 g of basic fuchsin in 87 ml of boiling
water. Cool to 50 C and add 30 ml of 1 M hydrochloric
acid and 3 g of potassium metabisulfite. Leave in the
dark for 24 hours to bleach. Add 0.5 g of decolourising
charcoal, shake and filter into a stoppered bottle. Store
in a cool, dark place.
For potassium metabisulfite,
see Hazcard 97. For
hydrochloric acid , see
Hazcard 47A. Wear eye
protection. Use a fume
cupboard.
No hazard label is required.
Leishman's
stain for blood
cells
Dissolve 0.15 g of the solid stain in 100 ml of
methanol.
For methanol, see Hazcard 40.
Prepare the solution in a fume
cupboard wearing eye
protection and disposable
nitrile gloves.
Label the solution TOXIC and
HIGHLY FLAMMABLE.
Methyl green
pyronin for DNA
and RNA
1 g of methyl green pyronin is dissolved in 100 ml of
distilled/deionised water. DNA is stained green and
RNA is stained red.
Methylene blue
for cheek cells
Wear eye protection, and gloves to avoid staining the
skin. Dissolve 0.1 to 1 g in 100 ml of water.
For methylene blue, see
Hazcard 32. The solution is
low hazard.
Orcein acetic
(Acetic orcein)
for
chromosomes
The stock solution contains 2.2 g of orcein dissolved in
100 ml of glacial ethanoic acid. Dilute 10 ml of this
solution with 12 ml of water before use. This diluted
solution does not keep.
Hazcard 38. Wear goggles and
chemical-resistant gloves.
Label the concentrated
solution CORROSIVE.
Sudan III or IV
solutions for
lipids
Dissolve 0.5 g of dye in 70 ml of ethanol and 30 ml of
water, using a warm water bath, and filter.
Ethanol is HIGHLY FLAMMABLE
(see Hazcards 32 and 40).
Label the solution HIGHLY
FLAMMABLE. Wear eye
protection.

CLEAPSS Recipe Book
98 CLEAPSS 2011
90 Stains for electrophoresis
Methylene blue is less sensitive than Azure A and not so convenient to use in electrophoresis but it is
cheaper and adequate for some activities.

Azure A for DNA Dissolve 0.04 g in 100 ml of 20% ethanol (ie, 0.04%
w/v) to apply after running the gel. Destaining is not
needed.
Low hazard.
Colloidal
Coomassie Blue
for proteins
See
www.ncbe.reading.ac.uk/NCBE/SCIENCEYEAR/recipes.html
for a recipe, or purchase from this organisation. Note
that this stain needs to be made up at least 24 hours
before use, and requires the addition of sodium
chloride solution to stabilise it.
Low hazard.
Methylene blue
for DNA
Dissolve 0.05 g in 200 ml water (ie, make up to
0.025% w/v) to apply after running the gel. Destaining
is needed.
See Hazcard 32, The solution
is low hazard.

CLEAPSS Recipe Book
CLEAPSS 2011 99
91 Stains for fungal material
Lactophenol Dissolve 10 g of phenol in 10 ml of water (do not heat).
Add 10 ml of propan-1,2,3-triol (glycerol) and 10 ml
lactic acid.
For propan-1,2,3-triol, see
Hazcard 38C. For phenol, see
Hazcard 70.Wear goggles.
Wear disposable nitrile gloves.
Use a fume cupboard. Label
the solution TOXIC and
CORROSIVE.
Cotton blue
(aniline blue) for
fungi
Dilute 8 ml of 0.1% cotton blue solution in lactophenol
and 4 ml of 0.1% basic fuchsin solution in lactophenol
with 88 ml of lactophenol.
See Hazcard 38C.
Fuchsin is HARMFUL. Wear
suitable eye protection and
gloves. Label the solution
TOXIC.

CLEAPSS Recipe Book
100 CLEAPSS 2011
92 Stains for metabolic activity
Janus green B
(Diazine green)
for mitochondria
Dissolve 0.3 g of the dye in 100 ml of purified water.
Dilute this solution ten times with water before use.
The colour change is blue (with oxygen) to pink
(anaerobic).
See Guidance Leaflet PS 89 for more information.
making up the solution. No
hazard label is required.
Methylene blue
for cell contents
For living organisms: dissolve 1 g solid in 100 ml water
and add 0.6 g sodium chloride. For dead tissue:
dissolve 0.23 g solid in 23 ml of ethanol and dilute to
100 ml with water.
For ethanol, see Hazcard 40.
For methyl blue see Hazcard
32. Wear eye protection and
No hazard label is required.
Neutral red Dissolve 0.1 g of neutral red in 100 ml of the
appropriate isotonic saline solution (see Recipe Sheet
75). Dilute the solution ten times further with the saline
solution.
Resazurin
solution to test
for milk
freshness
Dissolve one tablet or 0.005 g of resazurin powder in
50 ml of water. (1 ml of this is added to 10 ml of a
sample of milk. A colour change from blue to pink to
white indicates how many bacteria are present.)
Resazurin is IRRITANT.
TTC, to show
respiratory
activity
Dissolve 1 g of 2,3,5-triphenyl tetrazolium chloride
(TTC) in 100 ml of water. (A 0.5% solution is less
expensive and gives just as good results but takes
longer. It works well with maize seedlings.) Produces a
red colouration.
Low hazard.

CLEAPSS Recipe Book
CLEAPSS 2011 101
93 Stains for plant material
Aniline
(phenyl-
ammonium)
sulfate stain for
lignin
Mix 89 ml of ethanol, 10 ml of 0.05 M sulfuric acid and
1 g of phenylammonium sulfate [aniline sulfate(VI)].
For phenylammonium
sulfate(VI): see Hazcard 4.
making up the solution.
Label the stain HIGHLY
FLAMMABLE.
FABIL for plant
tissues
Prepare 3 solutions: 0.5 % solution of aniline blue in
lactophenol, 0.5% solution of basic fuchsin in
lactophenol and a solution containing 0.3 g of iodine
and 0.6 g of potassium iodide in 100 ml of lactophenol.
When required, mix in the proportions of 4:1:5 and
allow to stand overnight. Filter before use.
(Cell contents stain blue, cellulose walls stain light blue
and lignin stains yellow.)
For lactophenol, see
Hazcard 38C; for iodine, see
Hazcard 54.
Wear goggles and chemical-
resistant gloves.
Label the stain TOXIC.
Iodine stain Use 0.01 M iodine solution. See Recipe Sheet 50.
Phloroglucinol
for pentoses
and lignin
Dissolve 5 g of phloroglucinol (benzene-1,3,5-triol) in
75 ml of ethanol and 25 ml of water. Ligneous tissue
should be well-flooded and staining continued for
about 4 minutes after which 1 drop of concentrated
hydrochloric acid should be added.
Phloroglucinol is an IRRITANT.
Ethanol is HIGHLY FLAMMABLE
(see Hazcards 12, 40 and 47).
Label solution HIGHLY
FLAMMABLE. Wear eye
protection.
Schulzes
solution for
cellulose
Dissolve by warming 20 g of anhydrous zinc chloride in
8.5 ml of water and allow the mixture to cool. In a
separate container, dissolve 1 g of potassium iodide
and 0.5 g of iodine in 20 ml of water. Add this solution
dropwise to the zinc chloride solution until iodine
precipitate persists on agitation.
Zinc chloride is CORROSIVE and
iodine is HARMFUL (see
Hazcards 54 and 108). Wear
eye protection, and chemical-
resistant gloves, and carry out
the procedure in a fume
cupboard. Label the solution
CORROSIVE.

CLEAPSS Recipe Book
102 CLEAPSS 2011
94 Standard solutions for titration
A primary standard solution is one with a concentration that is accurately known. (They do not have to be
0.1 M) They are used to determine the concentration of other reagents undergoing titration.
Solids used should be of high purity (ie, more than 99.5% pure). Hydrated salts should not be used.
The solid should not alter composition during weighing (which is why sodium hydroxide is not used; it
absorbs water and carbon dioxide).
Distilled or deionised water must be used.
If possible, use a balance reading to 3 decimal places.
0.1 M sodium carbonate solution for standardising strong acid solutions

Procedure to prepare 1000 ml of solution
Heat the anhydrous sodium carbonate in an evaporating basin with a gentle flame for 30 minutes and
allow it to cool in a desiccator.
Measure 10.599 g of solid into a clean 250 ml beaker and add 150 ml of boiled pure water.
Stir the solution until the solid dissolves and pour it via a funnel into a 1000 ml volumetric flask.
Rinse the stirring rod and the beaker with water, pouring the washings into the volumetric flask.
Repeat this twice more.
Add water via the funnel into the flask so it is just up to the required mark. Add the stopper and mix the
solution well.
Titrate against a strong acid using methyl red as the indicator. The colour of the endpoint is orange.

0.1 M potassium hydrogenphthalate solution for standardising strong alkali
solutions

Measure 20.433 g of the solid into a clean 250 ml beaker and add 150 ml of boiled pure water.
solution well.
Titrate against a strong alkali using phenolphthalein as the indicator. The colour of the endpoint is pale
mauve which should last for at least 30 s.

0.1 M sodium ethanedioate solution for standardising potassium manganate(VII)
solution

Measure 13.40 g of solid into a clean 250 ml beaker. Add 150 ml of cold pure water. Stir the solution
until the solid dissolves and pour it via a funnel into a 1000 ml volumetric flask.
solution well.
Place the potassium manganate(VII) in the burette. Titrate against hot standard sodium ethanedioate
solution until there is a permanent mauve colour.
CLEAPSS Recipe Book
CLEAPSS 2011 103
0.0167 M potassium iodate(V) solution for standardising sodium thiosulfate solution

Dry the potassium iodate(V) at 180 C for 30 minutes before cooling and using.
Measure 3.574 g of solid into a clean 250 ml beaker and add 150 ml of cold pure water.
solution well.
To use, begin with a measured volume (10, 20 or 25 ml) of potassium iodate(V) solution. Add 10 ml of
1 M potassium iodide solution and 10 ml of 1 M sulfuric(VI) acid to produce iodine against which
sodium thiosulfate is titrated. Use a 1% starch solution as the indicator when the solution is pale
yellow. And from the results it is possible to calculate the concentration of the thiosulfate solution.

0.1 M calcium carbonate solution for standardising EDTA solution

Dry the precipitated calcium carbonate at 150 C for 1 hour.
Measure 10.00 g of cooled solid into a clean 250 ml conical flask and add enough 2 M hydrochloric
acid to dissolve it, placing a filter funnel over the flask to stop spray.
Make up this solution in a 1-litre volumetric flask.
CLEAPSS Recipe Book
104 CLEAPSS 2011
95 Strontium chloride
Strontium chloride hexahydrate will absorb water from the atmosphere.

Formula: SrCl
2
.6H
2
-1


Procedure for preparing 100 ml of 0.1 M strontium chloride solution
Dissolve 2.67 g of strontium chloride-6-water in 70 ml of water.

NB:
If a school has only strontium carbonate, the solution can be made by adding 1.48 g of the carbonate to
100 ml of 0.1 M hydrochloric acid.
CLEAPSS Recipe Book
CLEAPSS 2011 105
96 Sulfur dioxide
For acid rain see the Recipe sheet 97.
This is a particularly nasty gas to prepare and use. Do make sure that the procedure is trialled with help from
a more-experienced colleague and that the fume cupboard is working satisfactorily.

General Hazards See Hazcards 47A, 78, 92 & 97. Sulfur dioxide is TOXIC by inhalation. There is no
safe Workplace Exposure Limit for sulfur dioxide. As a guide, the HSE has advised
that levels, must be less than 2.7 mg m
-3
(ie, 1 ppm). Any preparation of sulfur
dioxide should be carried out in a fume cupboard.

Place between 10 and 20 g of sodium metabisulfite (HARMFUL) in a 250 cm
3
conical flask.
Set up the equipment as shown below.
Pour enough 2 M hydrochloric acid (IRRITANT) to just cover the bottom of the thistle funnel tube. Place
some moist acidified potassium dichromate(VI) paper (TOXIC) above the gas jar.
Heat the conical flask gently and collect the gas by downward delivery. When the acidified potassium
dichromate(VI) paper turns green, the gas jar is full.
Add 5 M hydrochloric acid to the metabisulfite if more sulfur dioxide is required.
The leftover reagents can be poured down the fume cupboard sink with plenty of water.
It is possible to carry out the fountain experiment. A round-bottom flask is substituted for the gas jar.

Heat
Add at least 50 cm
3
of
Loose mineral
wool plug
Acidified potassium
dichromate(VI) paper
CLEAPSS Recipe Book
106 CLEAPSS 2011
97 Sulfur dioxide solution
Sulfur dioxide is a particularly nasty gas to prepare and use. Do make sure that the procedure is trialled with
help from a more-experienced colleague and that the fume cupboard is working satisfactorily.

General Hazards See Hazcards 47A, 78, 92 & 97. Sulfur dioxide is TOXIC by inhalation. There is no
safe Workplace Exposure Limit for sulfur dioxide. As a guide, the HSE has advised
that levels must be less than 2.7 mg m
-3
(ie, 1 ppm). Any preparation of sulfur
dioxide should be carried out in the fume cupboard.

Method 1
Use a fume cupboard. Make sure it is working Wear goggles.
Place between 10 and 20 g of sodium metabisulfite (HARMFUL) into a 250 cm
3
conical flask.
Set up the equipment as shown below. The inverted filter funnel should be held just below the surface
of the water in the beaker.
Pour enough 2 M hydrochloric acid (IRRITANT) to just cover the bottom of the thistle funnel tube in the
flask.
Heat the conical flask gently to produce the gas, which will dissolve as it meets the water in the
beaker. The inverted funnel prevents suck back.
Add 5 M hydrochloric acid if more sulfur dioxide is required.
The leftover reagents can be poured down the fume cupboard sink with plenty of water.

Method 2
In a fume cupboard, dissolve 9.5 g of sodium metabisulfite (HARMFUL) in 100 ml of water. Add 100 ml of
0.5 M sulfuric(VI) acid and make up to 250 ml.

Acid Rain
Pupils should open containers in a fume cupboard and particular attention should be paid to known
asthmatics.
For acid rain, use 0.0001 M sulfuric(VI) acid rather than a sulphur dioxide solution.
For an acidic atmosphere, use a 0.1 M solution of sodium metabisulfite in water. This will generate enough
SO
2
gas to provide an acid atmosphere.
Heat
Add at least 50 cm
3
of
Inverted funnel
just below water
surface
CLEAPSS Recipe Book
CLEAPSS 2011 107
98 Sulfuric(VI) acid
The procedure below uses ice made from distilled or deionised water. Make sure you have a supply of this in
the freezer.
Do not make dilute solutions for the first time without seeking practical advice from a more-experienced
colleague.

Formula: H
2
SO
4
-1

General Hazards See Hazcard 98A. Addition of acid to water produces a lot of heat. NEVER ADD
WATER TO THE ACID; serious accidents have occurred when this has been done.

Concentration
required
label
500 1000 2500
0.01 M Ten-fold dilution of a 0.1 M solution with water -
0.1 M Ten-fold dilution of a 1 M solution with water -
0.4 M 11 22 54 -
1 M 27 54 136 IRRITANT
Battery acid 115 230 575 CORROSIVE
5M 135 270 680 CORROSIVE

Procedure
Wear goggles (a face shield is preferable when handling large volumes) and chemical-resistant
gloves.
Measure out the indicated volume of concentrated sulfuric(VI) acid in a measuring cylinder.
Fill the beaker or laboratory jug half to two thirds full with ice, add 200 ml of water and a stirrer bar.
Set the stirrer running on a magnetic stirrer and add the concentrated sulfuric(VI) acid slowly onto the
ice.
Keep stirring the solution until the ice melts.
CLEAPSS Recipe Book
108 CLEAPSS 2011
99 Testing for gases
Ammonia
Use damp red litmus
paper
Damp red litmus goes blue. Universal indicator can
be used as well.
Place concentrated
hydrochloric acid close by
White smoke of ammonium chloride (IRRITANT) forms. Wear eye protection.
Carbon dioxide
Use lime water See Recipe Sheet 20 for calcium hydroxide. A white
precipitate of calcium carbonate forms which, in time,
goes colourless as calcium hydrogencarbonate
forms.
Wear eye protection if
blowing into limewater
through a straw.
Carbon monoxide
Light it in a fume
cupboard
The gas catches fire with a blue flame forming
carbon dioxide.
Chlorine
Use damp blue litmus
paper
Damp blue litmus goes red on formation of chloric(I)
acid. This then bleaches the paper.
Dinitrogen monoxide
Add a lighted or glowing
splint
The gas supports combustion and can relight a
glowing splint, although it is not as active as oxygen.
Hydrogen
Add a lighted splint If the hydrogen is mixed with air there is a pop but if
the gas is pure it catches fire and there is no pop.
Hydrogen chloride
Use damp blue litmus
paper
Damp blue litmus goes red. Universal indicator can
be used as well.
Place concentrated
ammonia solution close
by
White smoke of ammonium chloride forms. Wear eye protection.
Hydrogen sulfide
Moisten filter paper with
0.05 M lead(II) nitrate(V)
solution
The paper immediately turns black with the formation
of lead sulfide.
Use tongs or forceps to
hold the paper.
Nitrogen
There is no positive test for nitrogen.
Nitrogen dioxide
Observe the colour The only brown gas there is. Wear eye protection.
Nitrogen monoxide
Add the gas to air Brown nitrogen dioxide is formed. Wear eye protection.
Oxygen
Add a glowing splint This relights a glowing splint, sometimes with a pop. Wear eye protection.
Sulfur dioxide
Moisten filter paper with
0.05 M acidified
potassium dichromate(VI)
solution
The paper goes green with the formation of
chromium(III) ions.
CLEAPSS Recipe Book
CLEAPSS 2011 109
100 Testing for negative ions
Bromide ions
0.05 M silver nitrate(V)
solution
Dissolve 0.85 g of silver nitrate(V) in 100 ml of
distilled/deionised water. A cream precipitate forms which
is soluble after the addition of concentrated ammonia.
See Hazcard 6 & 87.
Carbonate ions
2 M hydrochloric acid Adding the acid produces a lot of fizzing. The carbon
dioxide should be identifiable with lime water.
See Hazcard 47A.
Chloride ions
solution
Dissolve 0.85 g of silver nitrate(V) in 100 ml of water,
which must be pure. A white precipitate forms which is
soluble after the addition of 2 M ammonia solution.
See Hazcard 6 & 87.
Ethanoate and methanoate ions
0.1 M iron(III) chloride
solution
Add 2 M ammonia solution to 0.1 M iron(III) chloride
solution until a precipitate just appears. Now add more of
the original iron(III) chloride in drops with stirring until the
precipitate just dissolves. Addition of this neutral iron(III)
chloride to both ethanoates and methanoates gives a red
solution.
See Hazcard 6 & 55C
Iodide ions
solution
Dissolve 0.85 g of silver nitrate(V) in 100 ml of
distilled/deionised water. A yellow precipitate forms which
is not soluble after the addition 2 M ammonia solution.
See Hazcard 6 & 87.
Nitrate(III) (nitrite) ions
2 M hydrochloric acid Adding the acid produces a pale blue liquid and a brown
gas (nitrogen dioxide, TOXIC). Immediately, add water to
quench the reaction.
See Hazcard 47A, 68
and 93. Wear eye
protection.
Nitrate(V) ions
Brown ring test Dissolve the nitrate(V) in about 1 ml of water and add
0.5 g of iron(II) sulfate(VI)-7-water. Hold the test tube at
an angle and carefully pour about 1 ml of concentrated
sulfuric(VI) acid down the side of the test tube so it forms
a layer at the bottom. A brown ring indicates nitrate(V)
ions present.
Wear goggles.
See Hazcards 55B &
98.
Devadas alloy Add 2 ml of suspected nitrate(V) to 5 ml of 1 M sodium
hydroxide solution. Add a little Devardas alloy (contains
aluminium, copper and zinc) and warm. Test for ammonia
gas.
Wear goggles.
See Hazcard 1 & 91.
Phosphate ions
0.5 M ammonium
molybdate(VI) solution
(HARMFUL)
Dissolve 4 g of ammonium molybdate(VI) in 4 ml of
concentrated ammonia and 6 ml of water. Add 12 g of
ammonium nitrate(V) and dilute to 100 ml. When testing a
sample, acidify the sample first with 0.5 ml of 1 M nitric(V)
acid and then add 2 ml of the ammonium molybdate(VI)
solution.
See Hazcards 6, 8, 9A
& 67.
Sulfate(VI) ions
0.1 M barium chloride
solution (HARMFUL)
Dissolve 2.44 g in 100 ml of distilled/deionised water. A
white precipitate is formed. Sulfate(IV) ions do not give a
precipitate but it can form in time due to oxidation by
dissolved oxygen.
See Hazcard 10A.
CLEAPSS Recipe Book
110 CLEAPSS 2011
101 Testing for positive ions
Metal ions in general
Reagent Recipe and observations Hazards and notes
1 Flame tests Barium ions Greenish flame
Calcium ions Brick red flame
Copper(II) ions Bluish flame
Lead(II) ions White flame
Lithium Crimson red flame
Potassium ions Lilac flame (best seen thro cobalt glass)
Rubidium ions Blue flame
Strontium ions Crimson flame
Sodium ions Persistent yellow flame.
See Hazcards 10A,
19A & 47B.
The Hazcards also
have the procedure for
the tests. There is also
the borax bead test on
Hazcard 14.
2 Add 0.4 M
sodium
hydroxide to a
small amount of
the solution
under test
Dissolve 1.6 g of sodium hydroxide in 100 ml of water.
Precipitates form, some coloured and others colourless,
some of which dissolve in excess alkali solution.
Aluminium ions White precipitate which dissolves in
excess alkali
Ammonium ions No precipitate but odour of ammonia
Barium ions White precipitate
Calcium ions White precipitate
Cobalt(II) ions Blue precipitate
Copper(II) ions Blue precipitate
Iron(II) ions Green precipitate
Iron(III) ions Brown precipitate
Lead(II) ions White precipitate which dissolves in
excess alkali
Magnesium ions White precipitate
Nickel(II) ions Green precipitate
Silver ions Brown precipitate
Zinc ions White precipitate which dissolves in
excess alkali
Potassium ions No precipitate
Sodium ions No precipitate
See Hazcard 91.
The concentration of
the metal solutions
needs to be 0.1 M.
3 0.1 M sodium,
potassium or
ammonium
sulphide
solution
Dissolve 2.4 g of sodium sulfide-9-water or 1.0 g of
potassium sulfide in 100 ml of water. 4 ml of ammonium
sulfide solution can be diluted to 100 ml with water. Some
metal ions form precipitates with the solutions. Refer to an
(old!) A-level text for details.
See Hazcard 51. The
test solutions are
IRRITANT. Use a fume
cupboard as the odour
is repulsive.
Aluminium ions
Alizarin solution Dissolve 0.1 g of alizarin in 100 ml of water. Addition of 2 M
ammonia solution to an aluminium salt in the presence of
alizarin gives a pink precipitate or lake.
See Hazcard 32.
Ammonium ions
0.1 M sodium
hexanitrocobalt-ate(III)
solution (HARMFUL)
Dissolve 4.04 g in 100 ml of water. A yellow precipitate
forms with ammonium ions.
See Hazcard 95A. It is
an unstable solution
with a maximum shelf
life of 3 weeks.
0.4 M sodium
hydroxide solution
(IRRITANT)
Warm with the test solution. Ammonia gas will be given
off.
Calcium ions
0.25 M ammonium
ethanedioate
Dissolve 3.5 g of ammonium ethanedioate in 100 ml of
water. Ammonium ethanedioate gives a white precipitate
of calcium ethanedioate. The precipitate can be filtered off
and dried. It decomposes under heat to give calcium
oxide. The oxide can be weighed to estimate
quantitatively the amount of calcium present in a sample.
See Hazcard 36A.
CLEAPSS Recipe Book
CLEAPSS 2011 111
Cobalt(II) ions
Thiooxamide
(rubeanic acid)
(HIGHLY FLAMMABLE,
HARMFUL)
Dissolve 0.5 g in 100 ml of ethanol. A brown precipitate is
produced with cobalt ions.
See Hazcards 35 and
also 40A.
0.1 M ammonium or
potassium thiocyanate
Dissolve 0.76 g of ammonium thiocyanate or 0.97 g of
potassium thiocyanate in 50 ml of water and make up to
100 ml. A blue solution forms with cobalt(II) ions.
See Hazcard 9B or
95C.
Copper(II) ions
2M ammonia solution Dilute 11 ml of 880 ammonia to 100 ml with water. A blue
precipitate is formed which dissolves in excess ammonia
solution to form a deep blue solution.
See Hazcards 6.
Prepare in a fume
cupboard.
Thiooxamide
(rubeanic acid)
(HIGHLY FLAMMABLE,
HARMFUL)
Dissolve 0.5 g in 100 ml of ethanol. A black precipitate
forms with copper(II) ions.
See Hazcards 35 and
also 40A.
0.1 M ammonium or
potassium thiocyanate in water 50 ml of water and make
up to 100 ml. A black precipitate forms.
See Hazcard 9B or
95C. The solution is
LOW HAZARD.
Iron(II) ions
0.1 M potassium
hexacyanoferrate(III)
Dissolve 3.29 g in 100 ml of water. A light blue precipitate
forms.
See Hazcard 79. No
hazard warning is
required on the
solution.
1,10 phenanthroline
solution
Dissolve 1.49 g of 1,10 phenanthroline in 100 ml of water.
The reagent gives a red colour when mixed with iron (III)
ions.
See Hazcard 55B.
Solid Phenanthroline is
TOXIC if swallowed. No
hazard warning is
required on the
solution.
Iron(III) ions
0.1 M potassium
Dissolve 3.29 g in 100 ml of water. A dark blue precipitate
forms.
See Hazcard 79. No
hazard warning is
required on the
solution.
Ferroxyl indicator
(HIGHLY FLAMMABLE)
Dissolve 2 g of sodium chloride, 0.1 g potassium
hexacyanoferrate(III) and 1.0 g of phenolphthalein in
50 ml of ethanol, and mix with 100 ml of water. Very
useful for detecting the rusting process.
See Hazcards 32, 40A
and 79.
0.1 M ammonium or
potassium thiocyanate in 50 ml of water and make up to
100 ml. A dark red solution forms. [NB there is no
colouration with iron(II) ions]
See Hazcard 9B or
95C. The solution is
LOW HAZARD.
Lead(II) ions
Dithizone Dissolve 0.005 g in 100 ml of dichloromethane or
methylbenzene. The reagent gives a red colouration with
lead(II) ions.
See Hazcards 28, 32
and 46. Label the
solution according to
which solvent is used.
Potassium iodide Prepare a 0.1 M solution. A bright yellow precipitate
forms.
See Hazcard 47B.

CLEAPSS Recipe Book
112 CLEAPSS 2011
Nickel(II) ions
Thiooxamide
(rubeanic acid)
(HIGHLY FLAMMABLE,
HARMFUL)
Dissolve 0.5 g in 100 ml of ethanol. A violet precipitate
forms.
See Hazcards 35 and
also 40A.
Butanedione dioxime
(dimethylglyoxime)
(HIGHLY FLAMMABLE &
HARMFUL)
Dissolve 1 g in 100 ml of ethanol. A red precipitate forms. See Hazcard 53.
Potassium ions
0.1 M Sodium
hexanitrocobalt-ate(III)
(HARMFUL)
Dissolve 4.04 g in 100 ml of water. A yellow precipitate
forms.
See Hazcard 95A. It is
an unstable solution
with a maximum shelf
life of 3 weeks.
Zinc(II) ions
2M ammonia solution Dilute 11 ml of 880 ammonia to 100 ml with water. A white
precipitate forms, which dissolves in excess ammonia
solution to form a colourless solution.
See Hazcards 6.
Prepare in a fume
cupboard. The solution
is LOW HAZARD.

CLEAPSS Recipe Book
CLEAPSS 2011 113
102 Testing for organic functional groups
Alcohols - primary, secondary & tertiary
Lucas reagent
(CORROSIVE)
Dissolve 68 g of anhydrous zinc chloride in 43 g of concentrated
(36%) hydrochloric acid. The time taken for turbidity to appear is
used to differentiate between the three classes of alcohols:
no visible reaction: primary alcohol,
solution turns cloudy in 3 - 5 minutes: secondary alcohol,
solution turns cloudy immediately, and/or phases separate:
tertiary or benzyl alcohol.
See Hazcards
47A & 108A.
Potassium
dichromate(VI) test
Identifies primary and secondary alcohols. To five drops of the
alcohol, add 10 drops of 1 M sulfuric(VI) acid and 4 drops of 0.1 M
potassium dichromate(VI) solution. Place the solution in boiling
hot water from a kettle. The colour change is orange to green.
See Hazcard 78 &
98A.
Potassium
manganate(VII)
test
Identifies primary and secondary alcohols. To five drops of the
alcohol, add 10 drops of 1 M sulfuric(VI) acid and 4 drops of
0.002 M potassium manganate(VII) solution. Place the solution in
boiling hot water from a kettle. The colour change is purple to
colourless.
See Hazcard 81 &
98A.

Aldehydes and ketones
Hydrogen sulfite
test
Dissolve 20 g of sodium metabisulfite in 100 ml of hot water.
White precipitate appears with the reagent is added to a carbonyl
compound.
See Hazcard 92.
Tollens reagent It does not work with ketones. This reagent is ammoniacal silver
nitrate(V), it must be made immediately before use and not stored.
The breakdown products are EXPLOSIVE. Add one drop of 0.4 M
sodium hydroxide solution to 1 ml of 0.1 M silver nitrate(V). Add
1 M ammonia solution until the precipitate just redissolves. On
warming with an aldehyde, a silver mirror forms.
See Hazcard 87.
Immediately
dispose of liquid
after use down
the foul-water
drain.
Schiffs reagent It does not work quickly with ketones. Dissolve 0.1 g of fuchsin in
100 ml of water and add 0.9 g of sodium metabisulfite followed by
2 ml of 2 M nitric(V) acid, so it is colourless. Addition of an
aldehyde restores the colour. Be aware that sulfur dioxide, a TOXIC
gas, may be evolved.
See Hazcards 32,
67 & 92.
Wear eye
protection.
The iodoform test Works with methyl ketones (CH
3
CO-), methyl secondary alcohols,
ethanol and ethanal. To fives drops of the liquid under test, add 5
drops of 0.01 M iodine, followed by enough 1 M sodium hydroxide
solution until the brown colour is removed. Warm in boiling water
bath and look for a yellow precipitate of iodoform.
See Hazcards
54A & 91.

Carboxylic acids
Neutral iron(III)
chloride
Add 2 M ammonia solution to 0.1 M iron(III) chloride solution until
a precipitate just appears. Now add more of the original iron(III)
chloride in drops with stirring until the precipitate just dissolves.
Add this to the liquid under test, and look for a red solution.
[Methanoic acid reacts with acidified potassium manganate(VII)
and potassium dichromate(VI) in a similar manner to alcohols.]
See Hazcards 6 &
55C.

CLEAPSS Recipe Book
114 CLEAPSS 2011
Double bonds or unsaturation
Bromine water Add 0.002 M bromine water which will go clear. See Hazcard 15B.
Potassium
manganate(VII)
(acidified)
Add an acidified 0.002 M solution of potassium manganate(VII)
which will go clear. If it goes brown, it is not acidified enough.
See Hazcards 81
and 98.
Potassium
manganate(VII)
(alkaline)
Alkaline potassium manganate(VII) solution is not stable so it
must be made in situ. Add enough solid anhydrous sodium
carbonate to make the acidified solution alkaline. Add alkaline
0.002 M solution of potassium manganate(VII) which should go
green.
See Hazcards 81
and 98.
OH groups in alcohols and acids
Phosphorus
pentachloride
To about 2 ml of the liquid or solid under test, add phosphorus
pentachloride with a small spatula. There is a vigorous reaction
and hydrogen chloride is produced.
See Hazcard 74.
Phenol
Neutral iron(III)
chloride
Add 2 M ammonia solution to 0.1 M iron(III) chloride solution until
a precipitate just appears. Now add more of the original iron(III)
chloride in drops with stirring until the precipitate just dissolves.
Add this to the material under test, and look for a purple
colouration.
See Hazcards 6 &
55C.

CLEAPSS Recipe Book
CLEAPSS 2011 115
103 Tin(II) chloride
Also known as stannous chloride.
Solutions stronger than 0.2 M will be difficult to prepare. This reagent has to be dissolved in concentrated
hydrochloric acid before diluting further with water. The final solution will be 1 M with respect to hydrochloric
acid.
The solution may be cloudy, so either leave it for some hours before decanting off the clear solution or filter
through fine filter paper.

Formula: SnCl
2
.2H
2
0 Molar mass: 225.63 g mol
-1

General Hazards See Hazcards 102 & 47A.

Concentration
required
label
100 250 1000
0.1 M 2.26 5.64 22.56 -

Procedure
Wear goggles and chemical-resistant gloves. Use a fume cupboard.
Measure out the indicated quantity of tin(II) chloride-2-water.
Add the solid to about 10% of the final volume of concentrated hydrochloric acid in a beaker.
Stir to dissolve. Do not warm the solution.
Pour the solution into a measuring cylinder or laboratory jug and dilute to the final volume with
distilled/deionised water.
Leave to clear.
Pour the solution into a labelled bottle and mix well.

CLEAPSS Recipe Book
116 CLEAPSS 2011
104 Water (sea and hard)
It is possible to obtain sea-water salt from several suppliers.
For acid rain, see Recipe Sheet 97.

A. Sea water
Dissolve the following salts in 250 ml of pure water and dilute to 1000 ml.
23.99 g of sodium chloride
0.74 g of potassium chloride
2.24 g of calcium chloride-6-water
10.89 g of magnesium chloride-6-water
4.01 g of anhydrous sodium sulfate(VI) [OR 9.10 g of sodium sulfate(VI)-10-water]
0.20 g of sodium hydrogencarbonate
0.09 g of sodium bromide
0.03 g of boric acid (if required)
0.01 g of strontium chloride (if required)

B. Permanent hard water
Measure 0.45 g of calcium sulfate(VI)-2-water and add it to 1000 ml of water in a bottle and leave it
overnight to dissolve.

C. Temporary hard water
Filter 130 ml of freshly-made calcium hydroxide solution (see Recipe Sheet 20).
Pass carbon dioxide though the solution so that calcium carbonate first precipitates and then, with
further carbon dioxide, re-dissolves to form calcium hydrogencarbonate. This may take some time.
Dilute this clear solution to 1000 ml.
CLEAPSS Recipe Book
CLEAPSS 2011 117
105 Winklers method for dissolved oxygen
This technique is better than other methods (eg, using phenosafranine) because the oxygen in the water is
fixed once the manganese(II) hydroxide is formed. However, the solutions are hazardous to prepare and
use, and should be added to the water sample as soon as possible.

General Hazards See Hazcards 60, 91 & 98. Great care must be taken in making the 8 M sodium
hydroxide solution. Gloves and eye protection (a face shield is preferable) should be
used. The preparation is very slow and must not be rushed.

Preparation of the solutions used in Winklers method
100 ml of 8 M
sodium hydroxide
solution
Wear goggles and chemical-resistant gloves. Consider using a fume cupboard.
Add 8 g of sodium hydroxide pellets to 70 ml of water in a 250 ml beaker.
Once it has dissolved, cool the solution in an ice bath.
When the temperature of the solution has dropped to below room temperature
add another 8 g. Stir until it has dissolved and cool again.
Repeat this procedure until a total of 32 g of sodium hydroxide has been added.
Add the solution to a 100 ml measuring cylinder and add water to the 100 ml
mark.
Stir carefully into this solution 14 g of potassium iodide.
Pour the solution into a labelled bottle and, when cool, mix well. Attach a
CORROSIVE hazard warning.
The manganese(II)
solution
Dissolve 50 g of manganese(II) sulfate(VI)-4-water or 45 g of manganese(II)
chloride-4-water in 80 ml of pure water and dilute to 100 ml.
Pour it into a labelled bottle, mixing well. Add a HARMFUL hazard warning.
For the titration
Prepare 0.01 M sodium thiosulfate solution (see Recipe Sheet 87) which can be
standardised against potassium iodate solution (see Recipe Sheet 94) for
accurate work.
The indicator
Prepare 1% starch solution. See Recipe Sheet 49.

Winklers method for dissolved oxygen
Procedure for using these solutions
Wear goggles.
Collect 250 ml of water in a 250 ml stoppered bottle, ensuring there is no air trapped inside.
Use a 1 ml pipette (or marked teat pipette) to transfer 1 ml of the manganese(II) solution to the water
sample, injecting the liquid below the surface of the water, and taking care not to blow any more air
into the water.
Use a similar technique to transfer 1 ml of the alkaline potassium iodide solution to the water sample.
Replace the stopper on the bottle without trapping any air and agitate the bottle to mix the reagents.
Allow the contents to stand for 10 minutes.
Now the brown manganese(III) hydroxide has precipitated, the contents of the flask can be transferred
to a larger flask or bottle.
Add 1.5 ml of concentrated sulfuric(VI) acid which will dissolve the precipitate and liberate iodine.
Titrate this solution with the standardised sodium thiosulfate solution, adding 2 ml of starch solution
near the end-point. (1 mol of oxygen is equivalent to 4 mol of thiosulfate solution.)
CLEAPSS Recipe Book
118 CLEAPSS 2011
106 Zinc sulfate(VI)
On standing, zinc sulfate(VI)-7-water loses water of crystallisation slowly to the air and becomes powdery.
However, it is more convenient to make solutions from the hydrated salt than the anhydrous salt which is
also available from suppliers.
The anhydrous salt readily absorbs water from the atmosphere during storage.

Formula: ZnSO
4
.7H
2
-1

General Hazards Zinc sulfate(VI) is HARMFUL. See Hazcard 108B.


Concentration
required
label
100 250 1000
0.1 M 2.88 7.19 28.75 -
1.0 M 28.75 71.88 287.54 HARMFUL

Procedure
Measure out the indicated quantity of hydrated zinc sulfate(VI).
Either pour the solution from the beaker into an appropriate measuring cylinder and add water to the
required level or add water to the required level in the laboratory jug.
CLEAPSS Recipe Book
CLEAPSS 2011 119
Index

Name Recipe
Sheet

Acetic acid 39

Acetic alcohol 41

Acetic orcein 89

Aceto carmine 89

Acid atmosphere 97

Acid base indicators 46

Acid rain 97

Agar 1

Alcohol tests 102

Alcohol/water solutions 2

Alcoholic potassium
hydroxide
71

Alcoholic solutions 2

Aldehyde tests 102

Alginate beads 3

Alizarin solution 101

Alkaline pyrogallol 84

Alum 4

Aluminium chloride 4

Aluminium etching 38

Aluminium potassium
sulfate(VI)
4

Aluminium solutions 4

Amino acid
chromatography
26

Amino acid tests 13

Ammonia gas 5

Ammonia solution 6

Ammonia test 99

Ammonium cerium(IV)
sulfate(VI)
22

Ammonium chloride 7

Ammonium
ethanedioate
101

Ammonium hydroxide 6

Ammonium iron(III)
citrate
52

Ammonium iron(III)
sulfate(VI)
52

Ammonium sulfide 101

Ammonium thiocyanate 101

Ammonium vanadate 8

Amylase 37

Analgesics
chromatography
26

Andrews arrangement
for gas preparation
24

Name Recipe
Sheet

Aniline blue 91

Aniline sulfate(VI) 93

Animal tissue
preservative
74

Anion tests 100

Aspirator 20

Azo dyes 9

Azure A 90

Barium chloride 10

Barium diphenylamine-
4-sulfonate indicator
49

Barium nitrate(V) 10

Barium solutions 10

Basic agar 1

Belousov-Zhabotinski
reaction
63

Benedicts qualitative
reagent
11

Benedicts quantitative
reagent
12

Bile salts 37

Biological buffers 18

Biro ink
chromatography
26

Bismuth nitrate(V) 14

Biuret reagent 15

Bleach 81

Blue-bottle reaction 63

Blueprints 52

B-R reaction 63

Bradys reagent 33

Briggs-Rausher reaction 63

Brodies fluid 16

Bromide ion test 100

Bromine water 17

Bromocresol green 46

Bromophenol blue 46

Bubble mixture 79

Buffers 18

Butanedione dioxime 101

B-Z reaction 63

Calcium carbonate
(standard solution)
94

Calcium chloride 19

Calcium hydroxide 20

Calcium nitrate(V) 19
CLEAPSS Recipe Book
120 CLEAPSS 2011

Name Recipe
Sheet

Carbohydrate tests 13

Carbon dioxide 99

Carbon dioxide gas 21

Carbon dioxide indicator 48

Carbon monoxide
preparation
42

Carbon monoxide test 99

Carbonate ion test 100

Carbonyl group test 33

Carboxylic acid test 102

Cation tests 101

Cellular respiration 13

Cerium(IV) solutions 22

Cerium(IV) sulfate(VI) 22

Cheek cells 89

Chemical garden 86

Chemiluminesence 23

China blue agar 1

Chloride ion test 100

Chlorine gas
preparation
24

Chlorine test 99

Chlorine water 25

Chlorophyll
chromatography
26

Chromatography
solvents
26

Chrome alum 27

Chromium(III) chloride 27

Citric acid 28

Clarkes fluid 41

Clarkes soap solution 79

Clock reactions 29

Cobalt(II) chloride 30

Cobalt(II) nitrate(V) 30

Cobalt(II) thiocyanate 30

Coles reagent 13

Colloidal Coomassie
blue
90

Complexometric
indicators
49

Coomassie blue 90

Copper electroplating 36

Copper etching 38

Copper(II) chloride 31

Copper(II) nitrate(V) 31

Copper(II) solutions 31

Copper(II) sulfate(VI) 31

Name Recipe
Sheet

Cotton blue 91

Cotton dying 9

Cresol red 46

Crude oil alternative 32

Cytological fixative 41

DCPIP 13

Diammonium iron(II)
sulfate(VI)
51

Diazine green 92

Diazonium salts 9
2,6-
dichlorophenol indo-
phenol
13

Dichromate(VI) titration
indicators
49

Dimethylglyoxime 101

Dinitrogen monoxide
preparation
42

Dinitrogen monoxide
test
99
2,3,5-
Dinitrophenylhydrazine 33
3,5-
Dinitrosalicylic acid 34

phosphate(V)
67

phosphate(V) -3- water
67

Disappearing cross
reaction
29

Disinfectants 81

Disodium hydrogen
phosphate(V)
67

Disodium hydrogen
67

Dissolved oxygen 105

Dithizone 101
2,4-
DNP 33

DNS 34

DNSA 34

Double bond test 102

Drosophila food base 35

Dyes 9

Dying cotton 9

EDTA titration indicators 49

Ehrlichs haematoxylin 89

Eiosin indicator 49

Electrolytic agar 1

Enzymes 37

Enzymes immobilised 3

Eosin 89

Eriochrome black 49
CLEAPSS Recipe Book
CLEAPSS 2011 121

Name Recipe
Sheet

Etching solutions 38

Ethanoate ion test 100

Ethanoic acid 39

Ethanol/water mixtures 2

Ethanolic potassium
hydroxide
71

FAA 41, 74

FABIL 93

Fehling's solution 40

Ferroin indicator 49

Ferroxyl agar 1

Ferroxyl indicator 101

Feulgen's stain 89

Fixatives in biology 41

Flame tests 101

Fluorescein indicator 49

Formaldehyde 58

Formalin alcohol fixative 41

Formalin-aceto fixative 41

Formalin-aceto-alcohol 74

Formic acid 59

Glucose nutrient agar 1

Glucose quantitative
test
12

Glucose test 11, 34

Gram Stain 88

Harcourt-Essen reaction 29

Hard water 104

Home-made indicators 46

Hydrochloric acid 43

Hydrogen 99

Hydrogen carbonate
indicator
48

Hydrogen chloride
preparation
42

Hydrogen chloride test 99

Hydrogen gas 44

Hydrogen peroxide 45

Hydrogen peroxide/
potassium iodide
reaction
29

Hydrogen sulfide
preparation
42

Hydrogen sulfide test 99

Hydrogen-sulfite test 102
2-
hydroxy-1,2,3-
tricarboxylic acid
27

Immobilised enzymes 3

Name Recipe
Sheet

Indicator agar 1

Iodide ion test 100

Iodine solution 50

Iodine stain 93

Iodoform test 102

Iron etching 38

Iron(II) chloride 51

Iron(II) solutions 51

Iron(II) sulfate(VI) 51

Iron(III) chloride 52

Iron(III) ions and iodide
ion clock reaction
29

Iron(III) solutions 52

Iron(III) sulfate(VI) 52

Janus green B 13, 92

Kaiserlings
preservative
74

Ketone tests 102

Knops culture solution 66

Lactophenol 91

Landolt iodine reaction 29

Lead etching 38

Lead nitrate(V) 53

Leishman's stain 89

Lime water 20

Lipase 37

Lipstick
chromatography
26

Lithium chloride 54

Litmus 46

Locating agents 26

Lucas reagent 102

Lugols iodine 88

Luminol 23

Magnesium sulfate(VI) 55

Malt agar 1

Manganese(II)
sulfate(VI)
56

Manometric fluid 16

Mayonnaise agar 1

Mercury(I) nitrate(V) 57

Mercury(II) chloride 57

Metal ion
chromatography
26

Methanal solution 58

Methanoate ion test 100

Methanoic acid 59

Methanol solution 2
CLEAPSS Recipe Book
122 CLEAPSS 2011

Name Recipe
Sheet

Methanolic potassium
hydroxide
71

Methyl benzoate
nitration
26

Methyl green pyronin 89

Methyl orange 46

Methyl orange
preparation
9

Methyl red 46

Methylene blue 13, 89, 90,
92

Milk agar 1

Millons reagent 13

Mohrs salt 51

Molischs solution 13

Murexide 49

Neutral Red 46, 92

Neutral iron(III) chloride 102

Nickel electroplating 36

Nickel(II) sulfate(VI) 60

Ninhydrin 13

Nitrate(III) ion test 100

Nitrate(V) test 100

Nitration of Methyl
benzoate
26

Nitric(V) acid 61

Nitrogen preparation 42

Nitrogen test 99

Nitrogen dioxide
preparation
42

Nitrogen dioxide test 99

nitrogen gas oscillating
reaction
63

Nitrogen monoxide
preparation
42

Nitrogen monoxide test 99

N-phenylanthranilic acid
indicator
49

Nutrient agar for
bacteria
1

Nylon rope experiment 62

OH group test 102

Orange II preparation 9

Orcein acetic 90

Organic functional
group tests
102

Orthophosphoric acid 65

Oscillating reactions 63

Oxygen preparation 64

Oxygen test 99

Name Recipe
Sheet

Oxygen levels in water 105

PAS reaction 13

Pepsin 37

Periodic acid Schiff
reaction
13
Permanently hard water 104

pH buffers 18

pH probe calibration
buffers
18

Phenanthroline solution 101

Phenol red 46

phenol-
indodichlorophenol
13

Phenolphthalein 46

Phenolphthalein
indicator agar
1

Phenylammonium
sulfate(VI)
93

Phenylthiocarbamide 13

Phenylthiourea 13

Phloroglucinol 93

Phosphate(V) ion test 100

Phosphoric(V) acid 65

Phosphorus
pentachloride
102

Physiological saline
solutions
75

Plant mineral
requirement solutions
66

Plant tissue
preservative
74

Polypeptide tests 13

Potassium chloride 68

Potassium chromate 69

Potassium
dichromate(VI)
70

Potassium dihydrogen
phosphate(V)
67

Potassium
101

Potassium hydrogen
phthalate (standard
solution)
94

Potassium hydroxide 71

Potassium iodate
(standard solution)
94

Potassium
iodate/sodium
metabisulfite reaction
29

Potassium iodide 72, 101

Potassium
iodide/potassium
persulfate clock reaction
29
CLEAPSS Recipe Book
CLEAPSS 2011 123

Name Recipe
Sheet

Potassium
manganate(VII)
73

Potassium
phosphate(V)
67

Potassium sulfide 101

Potassium thiocyanate 101

Precipitation indicators 49

Preservatives for animal
tissues
74

Propanone solution 2

Protein tests 13

PTC 13

PTU 13

PVA 78

Pyrogallol 84

Red cabbage indicator 46

Redox indicators 49

Reducing sugar test 11, 12

Resazurin solution 93

Ringers solution 75

Rubeanic acid 101

Sachs culture solutions 66

Safranin counterstain 88

Sakaguchi test 13

Saliva 37

Sandells solution 76

Schiffs reagent 102

Schulzes solution 93

Screened methyl
orange
46

Sea water 104

Silicate gels 86

Silver electroplating 36

Silver nitrate(V) 77

Silver nitrate(V) titration
indicators
49

Slime 78

Soap 79

Sodium carbonate 80

Sodium carbonate
(standard solution)
94

Sodium chlorate(I) 81

Sodium chloride 82

Sodium dihydrogen
phosphate(V)
67

Sodium dihydrogen
67

Sodium diphenylamine-
4-sulfonate indicator
49

Name Recipe
Sheet

Sodium ethandioate
(standard solution)
94

Sodium ethanoate 83

Sodium
hexanitrocobaltate(III)
101

Sodium
hydrogencarbonate
84

Sodium hydroxide 85

Sodium hydroxide test 101

Sodium hypochlorate 81

Sodium phosphate(V) 67

Sodium phosphate(V)
-12- water
67

Sodium silicate 86

Sodium sulfide 101

Sodium
tauroglycocholate
37

Sodium thiosulfate 87

Solochrome black 49

Stains for cells 89

Stains for
electrophoresis
90

Stains for fungal
material
91

Stains for metabolic
activity
92

Stains for plant material 93

Standard solutions 94

Starch agar 1

Starch indicator 49

Starch malt agar 1

Sterilising conditions 1

Strontium choride 95

Sudan III or IV solutions 89

Sugar chromatography 26

Sulfate(IV) ion test 100

Sulfate(VI) ion test 100

Sulfur dioxide
preparation
96

Sulfur dioxide test 99

Sulfur dioxide solution 97

Sulfuric(VI) acid 98

Technical agar 1

Temporary hard water 104

Tests for gases 99

Tests for negative ions 100

Tests for positive ions 101

Thermochromic liquid 30

Thiooxamide 101
CLEAPSS Recipe Book
124 CLEAPSS 2011

Name Recipe
Sheet

Thiosulfate/acid
reaction
29

Thiosulfate/iodine
titration indicator
49

Thymol blue 46

Thymolphthalein 46

Tin(II) chloride 103

Tollens reagent 102
2,3,5-
Triphenyl tetrazolium
chloride
13

Trypsin 37

TTC 13

Universal buffer 18

Universal indicator 47

Name Recipe
Sheet

Unsaturation in organic
chemistry
102

Vitamin C test 13

Wanklyns soap solution 79

Washing soda 80

Water 104

Water solutions for
plants
66

Winklers method 105

Yamada indicator 47

Zinc electroplating 36

Zinc sulphate 106

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