Spectroscopy Measures light (radiation) absorbed by species in
solution. Some radiation is absorbed by ground state electrons in
atoms or molecules. Radiation NOT ABSORBED by the component being
analysed reaches a detector. This detector measures intensity of
radiation that is not absorbed by the sample. Absorbance readings
are related to concentration. Samples are generally diluted. Slide
2 Types of chemical analysis GC smaller, volatile hydrocarbons HPLC
larger macromolecules (proteins, drugs) Flame test presence of
metal (emission) AAS quantitative concentration of metal UV
coloured compounds IR Bend & stretching of bonds in molecules
(presence of different functional groups) NMR nuclear spin states
(exact order of atoms) Slide 3 IR Infrared Spectroscopy Expensive,
but simple are available at lower cost. Similar to UV
spectrophotometer but operates in IR region lower energy than UV
light IR radiation does not have enough energy to promote electrons
to higher level, but causes changes in covalent bond s in
molecules. The technique is based on the idea that different bonds
are of different lengths and therefore absorb IR radiation
differently. Difference in radiation between a reference cell and
the sample cell is recorded. Slide 4 Cells are made from NaCl or
KBr as glass and plastic absorb IR, Fast once sample is prepared
Gases, liquids & solids can be used Absorption spectroscopy
Qualitative, rarely quantitative Plot of transmittance against wave
number (spectra look like upside down UV spectra). Slide 5 IR
Transmittance: measures how much light has been transmitted. 100%
transmittance means no energy has been absorbed. Absorbance is the
intensity of light remaining after some has been absorbed. Used to
identify organic compounds, in particular functional groups, design
of new drugs, protein analysis, checking quality of wine products,
tea leaves that have a smell, forensic identification of oils,
fibres, paint flakes Slide 6 All IR Spectra has what's known as an
unique fingerprint region. This is generally only useful for
comparing an unknown spectra with that of a known substance IR
spectra. Note 900-1400cm -1 is called the fingerprint region. It is
generally hard to ID stretches within this region. It is the peaks
that appear after the fingerprint region that we are interested in.
The high absorbance readings at the larger wave numbers correspond
and can inform us pretty clearly of the function groups that are
present in the sample. It must be noted that symmetric molecules do
not absorb, cannot be ID. Molecules with stronger bonds absorb high
energy. Problems with IR Spectra: Some bands may overlap others.
Slide 7 IR Slide 8 Amines show a similar broad peak closer to 3500
cm -1 (due to N-H stretch). C=O group has an absorption peak
between 1670 and 1750 cm -1. This peak would appear for carboxylic
acids and esters. The O-H group shows a broad peak ~3000cm -1, this
may be seen in carboxylic acids and alcohols. Slide 9 Can you find
the functional group that is showing the maximum absorbance in the
following spectra? C=O 1670-1750 C=O 1670-1750 O-H 2500-3300 Broad!
Slide 10 Make sure you are using your data book! C-H 2850-3300 O-H
2500-3300 or N-H 3350-3500 *Cannot be O- H (acid) as no presence of
C=O! Slide 11 NMR Nuclear Magnetic Resonance Spectroscopy Expensive
Hazardous because of the strong magnetic field Operates in the
radio region Very fast (milliseconds) Qualitative Absorption
spectrum Spectra can be low resolution or high resolution Slide 12
How does NMR work? Protons, neutrons & electrons can spin if
provided with radiation of the correct wavelength. If there is a
even number of protons and neutrons in the nucleus (called
nucleons) no overall spin is exhibited. If there is a odd number of
nucleons there will be an overall spin. Atoms of this kind will
align either with or against a magnetic pole when placed in a
strong magnetic field. The atoms will act like a magnet when the
correct amount of radiation is absorbed. The atom will then flip to
a higher or lower energy state. This variation in energy is called
the chemical shift and is measurable in ppm. Slide 13 NMR Continued
A flip or chemical shift can only occur when some of the odd
nucleon atoms (e.g. 1H and 13C) absorb radiation in the radio
region. When nuclei return to their lower energy level, the
difference in radiation is released at a specific radio frequency.
The chemical shift is measured relative to an organic compound
called TMS< whose absorbance is plotted as zero on a NMR
spectra. Slide 14 LOW RESOLUTION PROTON NMR Tells us the type of
bond (using chemical shift value) Tells us the type of bond (using
chemical shift value) The number of peaks tell us how many
different H environments there are. The number of peaks tell us how
many different H environments there are. Height of peak tells us
how many H atoms in each environment (a higher peak means there is
a greater number of H nuclei in that particular environment).
Height of peak tells us how many H atoms in each environment (a
higher peak means there is a greater number of H nuclei in that
particular environment). The area under the peak is called the
integration value. The area under the peak is called the
integration value. Slide 15 HIGH RESOLUTION PROTON NMR Same info
can be gained from such spectra. Each peak is split into smaller
peaks (a number of peaks) The number of peak splits tells us the
number of H atoms the carbon adjacent to the group we are looking
at has. No splits = no neighbouring H atoms or may be a O-H or N-H
group. 3 peak splits = 2 H on next group. 4 peak splits = 3 H atoms
on next group. Slide 16 Each peak in a 13 C NMR Spectra represents
a difference Carbon environment. The chemical shift away from the
TMS reference is used to determine the nature of the carbon
environment The chemical shift values are greater than those for
proton NMR. Carbon NMR spectra show no splitting. Slide 17 How many
Carbon environments? How many Hydrogen environments? Answer: 2
Carbon Environments 2 Hydrogen Environments Answer: 5 Carbon
Environments 4 Hydrogen Environments Slide 18 Answer: 3 Carbon
Environments 3 Hydrogen Environments Answer: 3 Carbon Environments
ONLY 1 Hydrogen Environment *BOTH CH 3 groups are said to be
equivalent Answer: 3 Carbon Environments 3 Hydrogen Environments
Equivalent C and H atoms Slide 19 Mass Spectrometry (MS) Expensive,
but portable models are cheaper Very sensitive uses g samples
Qualitative and Quantitative Operates at low pressures Sample is
vaporised and high energy electrons knock off electrons of
molecules & cause fragmentation. The different species are
separated on passing though an electric field and a magnetic field.
Slide 20 Used to determine relative atomic mass on periodic table!
Also used to identify organic molecules, qualitative and
quantitative analysis of proteins, determine protein structure and
abundance and mass of isotopes. Slide 21 We can utilise MS to
confirm molecular mass and organic structures using the produced
fragmentation patterns. Samples are bombarded with high speed
electrons. Ions are formed & accelerated to high speeds into a
magnetic field. Lighter ions are deflected more than heavy ions. +2
charge are deflected more than +1 charged ions. A mass charge
ration (m/z or m/e) is produced and ALWAYS plotted on the
horizontal axis. Slide 22 More than one positive ion can be formed
(this is what we see as fragmentation patterns). In the first
ionisation, where one electron is knocked off, a molecular or
parent ion is formed. Once molecules form the parent (molecular
ion) they can continue to break down into smaller pieces or
molecules of lower molecular weight. It is important to remember
that only charged species will produce a line on a mass
spectrograph. The highest peak (also called base peak) is found for
the most stable fragment that is 100% absorbed. Common ions formed:
CH 3 + - mass/charge = 15 Also CH 2 Ch 3 +, OH - and COOH + Slide
23 From the mass spectra above: Q1. Which ion is most likely to be
the molecular (or parent) ion? Q2. Which ion is most abundant (has
the highest base peak)? MS Slide 24 Slide 25 Combining
Techniques
