Basic concepts and tools

of
Analytical Chemistry
L3

2hr

Outline

SI units
Measuring weights and volumes
Preparing a solution
Analytical reagents, chemicals
pH determination
Buffer solutions
Data analysis

SI: Units

Science has units

SI: abbreviated from the French ( Systme International dunits)
Derived from the Systme mtrique (metric system)
In 1790 the French Revolution implemented the metre and kilogram as standards of length
and mass

Then later it was refined and extended to

Length : meter (m)

Mass: kilogram (kg)
Time : second (s)
Electric current: ampere (A)
Thermodynamic temperature: kelvin (K)
Amount of substance: mole (mol)
Luminous intensity: candela (cd)

Only 3 countries have not officially adopted the metric system

Liberia, Birmanie, USA
(although in the US most scientific measurements are using the metric system)

Non SI units accepted with SI

Sometimes SI units are not practical
Non SI units accepted with SI
Minute, hour, day, hectare, litre, tonne, ..

Non SI units whose values must be obtained

Electron volt, dalton, dalton/unified atomic mass unit, Planck
constant, electron mass, ..

Other non-SI units

Bar, angstrm, nautical mile, decibel, ..

1 liter
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A word of English
French :
British English:
American English:

mtre, litre, gramme, kilogramme

metre, litre, gramme, kilogramme
meter, liter, gram, kilogram

Expressions
e.g. la7n: exempli gra+a, for example
i.e. la7n: id est, in other words,

Concentrations units
Concentration = mass / volume
e.g.
gram per liter; g/L = g. L-1
mg/L, ng/mL, g/mL

Molarity

1 mg/L = 1 g/mL
1 g/L = 1 ng/mL

1 mole per liter = 1 mol/L = 1 mol.L-1 = 1M

10-6 mol/L = 10-6 mol.L-1 = 1 M

Conversion M to ng/mL
If the molecular weight of the compound is MW
then 1 M = MW x ng/mL
e.g.
MW of compound X is 300 then 1M of compound X is 300 ng/mL

Mass vs weight
Mass and weight are the same in common language

But weight is a force

The pound
1 lb = 0.453 59237 kg
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Measuring weights and volumes

It is important to know the concentration
Concentration = mass/ volume

We have have to measure with accuracy mass and

volume
A balance is an analytical instrument that measures
weight
But it is calibrated with mass so it measures mass

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We need the right tool

Weighing scales
A balance has an useful range
We have to choose the correct scale
.1-5 kg

10- 150 kg

> 1000 kg

50 g 200 g 100 kg

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Analytical balance
To weigh mg and g amounts we need an analytical
balance
It is an analytical instrument
It should be
accurate
reproducible
sensitive

Should be regularly calibrated with standard masses

Has a door to prevent air draft
precision : 0.1 mg

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Measuring volumes
We have to choose the correct container

Accuracy and precision

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How to read the volume?

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Laboratory pipettes

Pasteur
Graduated (measuring) pipettes, burets
Volumetric
Mechanical electronic pipettes

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pipette
Pasteur pipettes
Named after Louis Pasteur
in glass or in plastic to deliver small volumes (no accuracy
intended)
Cheap enough to be considered disposable
But can be reused if no contamination possible

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Graduated and volumetric pipettes

Graduated pipettes
To deliver known volumes
In glass or polystyrene for sterile serology

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burette
Graduated glass cylinder to dispense liquid drop by drop
with accuracy

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Volumetric pipettes

Safety:
Never pipet with your mouth

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Mechanical/electronic pipettes
Fixed volume or adjustable
Need a disposable plastic cone

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Small volume syringe

For small volumes
In the L range

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uncertainty
No measurement is free from error
Error is introduced by
The limitations of instruments and measuring devices
The imperfection of human senses

In analytical chemistry the estimated degree of error in a

measurement is called the uncertainty of the
measurement and the reported values are only with
significant figures

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Significant figures
Numbers that describe the value without exaggerating its
accuracy
We report as significant all numbers knows with aboslute
certainty, plus one more digit that is inderstood to contain some
uncertainty
The uncertainty in the final digit is usually assumed to be 1

There are some rules

http://2012books.lardbucket.org/books/principles-of-generalchemistry-v1.0m/s05-09-essential-skills-1.html

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Last significant figure in weighing

The reading on the analytical balance is 3.2479 mg
How good is that number ?
Should we report it ?
We know that the balance is accurate

The precision of the balance is 0.1 mg. It means that it can detect a
change of 0.1 mg
The weight should be reported as 3.2 0.1 mg
Why ?
The last significant number is the first decimal: .1 mg
The numbers 4 7 9 are meaningless. The balance cannot detect changes of:
. 01, .001 and .0001 mg

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Significant figures in measuring volume

Which measuring apparatus do you take to deliver 9.7
mL as accurately as possible ?
Then with how many significant figures can you measure the
volume ?
Use the 10mL graduted cylinder
Accurate with 2 signicant gures

hVp://2012books.lardbucket.org/books/principles-of-
general-chemistry-v1.0m/s05-09-essen7al-skills-1.html
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Amount and concentration

1 mole of any substance is N molecules of that substance
Avogrado number N= 6.022 1023
It is the number of carbon atoms in 12 g of 12C
N unit: mol-1

In biochemistry we are usually calling the unified atomic mass unit a

dalton Da (from John Dalton)
kDa kilodalton: for high mass polymers: proteins and nucleic acids

The molecular weight of a compound is the sum of the masses of

the elements
It is the sum of the mass of each constituent
H2O = 2 H + O = Da
CH4 = C + 4 H = Da

Molecular weight calculator

http://www.lenntech.com/calculators/molecular/molecular-weight-calculator.htm

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Molarity
I mole of a molecule contains N molecules with a mass
expressed in g (MW)
1 mole of water is 18.02 g of water
1 mole of NaCl is 58.44 g of NaCl

Conversely the Molecular Weight gives the number of g in a

mole of substance
18.02 g of water contains
1 mole of H2O molecules
N (Avogadro number) water molecules

Concentration
mol/L = mol.L-1 = M
1M NaCl = 1 mol/L NaCl = 58.44 g/L
A 0.1M NaCl solution contains 5.844 g of NaCl per liter

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Prepararing a solution
It is one of the most important task of bioanalytical
chemistry
What is a solution ?
A solution is a chemical compound (or chemicals) dissolved in a
liquid
usually at a defined concentration

There are 3 components in a solution

The chemical(s)
The solvent
The concentration

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Chemicals
Bionalytical chemistry is usually about measuring trace
amounts/ concentration in a biological sample
so the chemicals should not add impurities in the system

A chemical compound of a known high standard of

purity has to be used (Analytical grade)
There are specifications for reference compounds
We have to know what are the impurties
i.e. glucose

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Chemicals for analytical work

References compounds must be as pure as possible
You have to know what is in the compounds
Racemate/ enantiomer ?
% pure ?
What are the impurities ?

An analytical chemical/reagent needs a Certicate of

analysis
You get the chemical that is not giving interference with
your analysis

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American Chemical Society

hVp://pubs.acs.org/
reagents/demo/

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solvent
A solvent is a liquid chemical
The solvent should not add impurities to the system
Water is a solvent
Water has to be as pure as possible

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Storage of chemicals
A chemical/reagent/solution has a finite shelf-life which
depends on:
Temperature:
At what temperature should I store the chemical?
Room temperature/ refrigerated/ frozen

Humidity
Some compounds are hygroscopic
You need to store them in a dessicator

Length of time
A chemical may degrade with time. It has a limit date

Light
A chemical may be light sensitive

Oxidation
A chemical may be subject to oxidation

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concentration
The concentration may be expressed in 3 different ways:
Mole per Liter: M (or mM or M, ..)
mol/L

mass per volume g/L

The isotonic NaCl 0.9 % is a
0.9 g/100mL of water
9 g/L

Volume per volume : vol/vol

When we mix 2 solvents the solution is usually expressed in vol/vol
Ex: a 40% (vol/vol) methanol/water used in chromatography

When we talk about a solution we must know

how it is prepared: M, mass/vol or vol/vol
and when it was prepared

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Preparing a solution
To prepare 500mL of a 0.1 M NaCl solution
How much do we need ?
0.1 x 0.5 mole of NaCl
0.1 x 0.5 mole of NaCl = 0.1 x 0.5 x MW(NaCl) in g
= 0.1 x 0.5 x 58.44 = 2.922 g of NaCl

We dissolve 2.922 g in 500 mL

We calculate the number of moles in the desired volume

to get the right concentration
The date of preparation should always be noted on the
container
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Tricks
Some compounds are present (crystallize) as:
Salts
Hydrates
Potassium phosphate tribasic anhydrous K3PO4
Potassium phosphate tribasic, octahydrate K3PO4, 8 H2O

So when we prepare a solution we have to know the physical form

Anhydrous, Hydrate, Free base, salt

Ex trehalose is present
Anhydrous : melting point 203C, molar mass: 342.296 g/mol
Dihydrate: melting point 97 C, molar mass: 378.33 g/mol
If we want a tetralose 1M trehalose solution we disolve in 1 L
342.296 g if we start with the anhydrous form
378.33 g if we start with the dihydrate

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Tricks: acidic and basic compounds

With a basic compound always check:
Base form
Morphine: C17H19NO3 MW= 285.34

Salt and what kind of salt

Morphine hydrochloride: C17H19NO3.HCl MW = 312.81
Sulfate pentahydrate: 2C17H19NO3.H2SO4.5H2O

With an acidic compound always check

Acid form
Salt and what kind of salt

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Dilution
We have a solution and we want to dilute it:
2 ways
Final amount is the same

the final volume is increased

Final volume is the same

The final amount is decreased

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Dilution
Final amount is the same
We have a 10M HCl solution and we want to prepare
200mL of a 0.5 M HCl solution
What volume should we take ?
c1V1 = c2V2: the amount is the same before and after

C1 concentration before dilution

C2 concentration after dilution
V1 volume before dilution ( unknown)
V2 volume after dilution (what we want)

V1 = (c2/c1). V2
V1 = (0.5/10) x 200 10 mL de HCl 10 M
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Dilution
the amount does not change
The amount does not change, the volume increases
10 fold dilution
1 vol starting solution + 9 vol solvent

20 fold dilution
1 vol starting solution + 19 vol solvent

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Serial dilution
the volume does not change
Serial dilution
Can be easily automated

V1 = (c2/c1). V2
Beware: the error in pipe7ng
propagates
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errors
We have seen that measurements are uncertain
volume 25 0.03 mL
Weight 10 0.1 mg
Absolute uncertainty vs relative uncertainty
For the 25 mL volumetric pipette the volume is 0.03 mL
0.03/ 25 = 0.0012 = 0.12%
It is a fixed volume

The balance weighs at 0.1 mg

If we weigh
1 mg there is 10% uncertainty
10 mg there is a 1% uncertainty
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Error in concentration
Concentration = mass / volume
The error (uncertainty) of the concentration is the sum of
the errors
(concentra7on) = (mass) + (volume)

Ex: mass = 100 1 (1%) Volume = 100 1 (1%)

both values between 99 and 101
The ratio may be between 99/101 and 101/99
0.98 and 102 = 100 2%
(more on that later)
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Measuring the pH of a solution

With a pH meter
With pH indicators

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pH meter
pH meter
Precise 0.1 or 0.01 pH unit
Measures the potential difference between the working electrode
(usually glass) and a reference electrode
There are many hand held pH-meters
It is calibrated against solutions of known pH values

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pH indicators
It is a chemical that changes color
in a specific pH range

It changes color in a pH interval

Bromocresol green
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phenophthalein

pH = 1

pH = 13

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pH indicators
test papers
The paper has been impregnated with dyes
The paper is dipped into the solution
the color is compared to a standard chart

Gives an indication

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Tutorials
Mole
http://chemcollective.org/activities/tutorials/stoich/the_mole

Dilution

Virtual lab
Can be downloaded
Chemcollective.org/vlabs

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