SPECTROSCOPY TYPES OF SPECTROSCOPY 1. Absorption Spectroscopy Passes the Radiation WaveA. UV-range=200-380nm B. Visible=380-780nmC. Near-IR=780-2500nmD. IR=2.5-40um

OtherNMR:Nuclear Magnetic ResonanceMass Spectroscopy

ULTRAVIOLET-VISIBLE SPECTROSCOPY•UV-range=200-380nm •Visible=380-780nm

It is used to measure the multiple bonds or atomic conjugation within the molecule. The UV-Visible region is subdivided as below

• Vacuum UV: 100-200 nm • Near UV: 200 to 400 nm• Visible region: 400 to 750 nm

1. Basics of UV Light Absorption:

UV Light Absorption Ultraviolet/visible spectroscopy involves the absorption of ultraviolet/visible light by a molecule causing the promotion of an electron from a ground electronic state to an excited electronic state Absorption of this relatively high-energy light causes electronic excitation.

The easily accessible part of this region (wavelengths of 200 to 800 nm) shows

absorption only if conjugated pi-electron systems are present.

2. Electromagnetic Spectrum

3.Electronic spectroscopy

When molecule absorb the energy , it results in electronic transition.

Electron promotion will be from the highest occupied molecular orbital (HOMO) to the lowest unoccupied molecular orbital (LUMO) and the resulting species is called an excited state.

Types of Transitions:

Types of Transitions There are several types of electronic transitions available to a molecule including:

• σ to σ * (alkanes) • σ to π * (carbonyl compounds) • π to π * (alkenes, carbonyl compounds, alkynes, Azo compounds)• ή to σ * (oxygen, nitrogen, sulfur, and halogen compounds) • ή to π * (carbonyl compounds)

4. Electronic transactions involved

σ * (anti-bonding)

σ * ή σ * σ E

π * (anti-bonding) ή π * π *

ή (non-bonding)

π π * π σ * π (bonding)

σ π * σ (bonding)

σ σ * Transitions:

σ σ * Transitions an electron in a bonding σ orbital is excited to the corresponding antI bonding orbital. The energy required is large.

For example, alkanes as methane (which has only C-H bonds, and can only undergo σ σ * transitions) shows an absorbance maximum at 125 nm. Absorption maxima .

Due to σ σ * transitions are not seen in typical UV-Visible spectra (200 -

700 nm). σ bonds are very strong and requires higher energy of vacuum UV.

ή σ * Transitions:ή σ * Transitions Saturated compounds containing atoms with lone pairs

(non-bonding electrons) are capable of ή σ * as transitions. These transitions usually need less energy than σ σ * transitions.

They can be initiated by light whose wavelength is in the range 150 - 250 nm. The number of organic functional groups with n σ * peaks in the UV region is small.

These transitions are involved in saturated compound with one hetero atom with unshared pair of electron i.e. saturated halides, ethers, aldehide, ketones, amines etc.

These transitions are sensitive to hydrogen bonding e.g. alcohol and ethers which absorbs at wavelength shorter than 185 nm therefore used as a solvent in UV.

π π * Transitions:

π π * Transitions For molecules that possess π bonding as in alkenes, alkynes, aromatics, acyl compounds or nitriles, energy that is available can promote electrons from a π Bonding molecular orbital to a π Antibonding molecular orbital ( π * ). This is called a π π * transition.

The absorption peaks for these transitions fall in an experimentally convenient region of the spectrum (200 - 700 nm).

These transitions need an unsaturated group in the molecule to provide

the π electrons.

ή π * Transitions:

ή π * Transitions Even lone pairs that exist on Oxygen atoms and Nitrogen atoms may be promoted from their non-bonding molecular orbital to a π antibonding molecular orbital within the molecule. This is called an ή π * transition

These are available in compounds with unsaturated centers. e.g. Alkenes. They requires lowest energy as compare to others.

Terminology

Chromaphore:

A covalently bonded unsaturated group responsible for electronic absorption.

These are group of atoms that absorbs light whether or not a color is thereby produced. Eg c=c, c=o, No2 etc

A compound containing chromaphore is called chromogen. There are two types of chromaphore .

Independent chromaphore: single chromaphore is sufficient to import color

to the compound e.g. Azo group Dependent chromaphore: When more than one chromaphore is required to

produce color. Eg acetone having 1 kentone group is colorless where as diacetyl having two kentone groups is yellow.

Auxochrome:

A saturated group with non bonding electron when attached to chromaphore alters both wavelengths as well as intensity of absorption. Eg OH, NH2, NHR etc.

A group which extends the conjugation of a chromaphore by sharing of

nonbonding electrons.

•

Bathochromic group: The group which deepens the colour of chromaphore is called bathochromic

group. Eg. Primary, secondary and tertiary amino groups.

Bathochromic shift: (Red shift) shift of lambda max to longer side or less energy is called bathochromic shift or read shift. This is due to substitution or solvent effect.

Hypsochromic shift:

(Blue shift) shift of lambda max to shorter side and higher energy is called hypsochromic or blue shift. Eg solvent effect. That may be increase or decrease absorption intensity.

SOME SPECTRUMS OF ORGANIC ANALYTES

Instrumentation:

A beam of light from a visible and/or UV light source (colored red) is separated into its component wavelengths by a prism or diffraction grating.

Each monochromatic (single wavelength) beam in turn is split into two equal intensity beams by a half-mirrored device.

One beam, the sample beam (colored magenta), passes through a small transparent container (cuvette) containing a solution of the compound being studied in a transparent solvent.

The other beam, the reference (colored blue), passes through an identical cuvette containing only the solvent.

The intensities of these light beams are then measured by electronic detectors and compared. The intensity of the reference beam, which should have suffered little or no light absorption, is defined as I0 . The intensity of the sample beam is defined as I . Over a short period of time, the spectrometer automatically scans all the component wavelengths.

Solvent effect: : Different compounds may have very different absorption maxima and

absorbances. Intensely absorbing compounds must be examined in dilute solution, so that

significant light energy is received by the detector, and this requires the use of completely transparent (non-absorbing) solvents.

The most commonly used solvents are water, ethanol, hexane and cyclohexane. Solvents having double or triple bonds, or heavy atoms (e.g. S, Br & I) are generally avoided.

Choice of Solvent:

First criteria for good solvent are that it does not absorb uv-radiation in the same region. As the substance whose o-absorptivity is not meaning that it does not absorb radiation.

Example: The solvent do not contain conjugated system H2O, ETHANOL (95%), HEXANE,.

Second criteria for absorption if it on fine structure on absorption band. Non-

polar solvent (not contain) do not make H-bond with the solute (organic solvent) and spectrum of solvent is approximate to gases state (steam is puriest form), in which fine structure often observe. In molar solvent the H-bonding forms a solute-solvent complex and fine structure may disappear.

It should contain polar system and conjugated form.

UV-Visible spectrum:

UV-Visible spectrum A spectrum is a plot of the intensity of energy detected versus the wavelength

In case of single band of wavelength, < 220nm usually indicate ή- σ * (amine NH3), alcohol, ether and thiols .

Exception: ή- π * (cyano group).it can identify because of its weak transition

absorbtivity 100 (ether, alcohols and amines).which is further identify by IR. A single band of low absorbtivity,10-100,at 250-360nm,indicate ή- π *

transition ,indicate(CN,C=O,COOH,COR,N=N,C=N,NO2).

Two bands of medium intensity, absorbtivity 1000-10.000 both with Lambda (λ) above 200nm indicates aromatic system.(polynuclear aromatic substance a 3rd band appear near 200nm.where it cannot be observe.

Bands of high intensity, absorbtivity, 10,000-20,000 appear above

210nm.Generally represent alpha-beta unsaturated ketones, dienes, and polyene.

Simple ketones,acids,esters,amides and other compounds containing both π system and unsaturated electron pairs shows two absorption.

Compounds that are highly colored, exhibit their spectra in visible region, unlikely to contain long chain conjugated system or polycyclic aromatic system chromaphore.

If an organic compound does not absorb uv-visible radiation, it means that the compounds do not contain conjugated double bonds.

If an organic compound absorbs uv-vis radiation, it means that the compound contains a carbonyl group or conjugated double bonds.for example, conjugated dienes, carbonyl compounds and aromatic compounds all absorb in the uv-vis region.

APPLICATIONS Qualitative and quantitative analysis. SOURCE; deuterium lamp, hydrogen lamp. VISIBLE –SPECTROSCOPY Tungstun tamp,

IR-SPECTROSCOPY• RANGE=2.5u-40um• SOURCE=Zarconium-oxide,• Rare earth oxide • , nearst and global.

IR- radiation is weak; it does supply sufficient energy for bonds in the molecules to vibrate by stretching or bending.

As the molecules is subjected to the individual wavelength in the 5000-667cm-1 range,

Since different bonds and functional groups at different wavelength, an infrared spectrum is used to determine the structure of organic molecules.

For example, carbon-carbon triple-bond is stronger than a carbon-carbon double bond and requires a shorter wavelength (greater energy) to stretch).The same consideration apply to carbon-oxygen and carbon-nitrogen bonds.

An infrared spectrum studied in two section

i. Functional group regionii. Fingerprint region

i.Functional group region

The area from 5000cm-1-1300cm-1 is called the functional group region. The bands in this region are particularly useful in determining the type of functional groups percentage in the molecule.

ii.Finger print region

The area from 1300cm-1 to 667cm-1 is called the fingerprint region. A peak by peak match of unknown spectrum with the spectrum of the suspected compound in

this region can be used, much like a fingerprint to confirm its identity.

IR-SPECTRUM

EXAMPLESMETHANOL

BUTANONE

ISOBUTYLAMINE

INSTRUMENTATIONThe instrument that determines the absorption IR spectrum of a compound is called IR spectrophotometer.

TYPES: there are two types.

i. Dispersive –IR spectrometerii. Fourier transform Ir spectrometer

Both of these instrument travel spectra of compound in the common range of 4000-400cm-1.

DIR-spectrometer :

The instrument produce a beam of IR radiation from a hot wire (IR source).The beam of by means of mirror divided into two parallel beam of equal intensity radiation,

•Beam sample analysis•Reference

Both beams can pass through monochromatic, which disperse each into continuous spectrum of frequencies or IR-light by beam chopper. Which is a rotating chamber that passes the two beams alternately through a diffracting gratting serve as prism which varies of frequency or range?

The thermo-couple detector senses the ratio between the sample and the reference that what beam has bean absorbed by the sample and which uneffective.

After the singnal from the detector is amplifies the recorder cause the resulting specterum of sample on chart.

The IR-chart is recorded as a frequency of the IR-radiation charges by the rotation of the diffraction gratting.

Disperive spectrum is recorded in frequency domain, It is common to plot frequency is live transmission.This is recorded as percentage transmitter from detector from record the ratio of intensity of two beam therefore.

%T=is/ir x100 Is=sample beam intensity Ir=intensity of reference

SAMPLE PREPARATION To obtain the IR-spectrum compound is dissolved in a solvent or diluents .This

solution is then will replace in sample beam by pure solvent beam in a identical beam.

The instrument automatically subtracts the spectrum of solvent from that of sample. The instrument also cancel the effect of IR spectrum NO2, CO2.If pure liquid is analyzed the sample is varyingly in a sample beam and nothing is inserted in a reference beam.

When the spectrum of liquid is obtain. The effects of atmospheric gas automatically cancel, since they are present in both beams.

FTIR-SPECTROSCOPY

The most common IR-spectrometer are design to consist of optical pathway, produce a pattern called interferofram,(broader instrument optical pathway)There is optical pathway to analyses IR interferogram .

The interferogram is a complex signal but it’s waves like pattern contain all frequency that make up the IR-spectrum.

Interferogram is plot of intensity vs. time, called time domain spectrum.

FTIR-SPECTRUM In chemical analysis, frequency domain spectrum is preferred over time domain

spectrum is a plot of intensity vs. frequency vs. time.

OPEARTION OR WORKING OF FTIR

Computer interferogram FTIR instrument operate in a single mode to obtain spectrum .first interferogram of the background being recorded by example (co2).

This background interferogram subjected to Fourier transformer which yield

a spectrum of background., then sample is placed into the beam spectrum is obtain resulting from Fourier transform of interferogram .

The spectra contain absorption of sample and background and now where automatical subtract spectrum of background from the sample which identical to dispersive spectrum.

PREPARATION OF SAMPLE

To determine the IR-spectrum of compound sample is placed in holder or cell as must be constructed from ionic substance (NACL), KBr.

Glass or plastic absorb Irradiation, so it should be not.

KBr plates are expensive then NACL, and useful in the range of 4000-400cm-1.

NACL is useful in the range 4000-659cm-1 and band with frequency less than this value will not be observe ,since only a few band appear bellow 650cm-1.NACL.

LIQUID A drop of liquid is placed between a pair of polished and NACL and KBR plot refer to as

salt plates, where plots are squeeze gently a this liquid film are formed between them. Spectrum of obtain is know next spectrum since solvent.

Salt plates are water soluble, so analyzing compound must be free from water; the pair of plate is inserted, into a holder, which fixed into a spectrometer.

SOLIDS

Following common methods to grounded sample mix with powder KBr melts and seal the compound into a matrix results in FTIR plot or disc should be transport, which can be inserted into holder in IR-spectrometer.

ADVANTAGE: In case of wood plates for disc the spectrum have no interfering band, since KBr transport down to 4000cm-1.

DISADVATNTAGE: KBr through absorb water which may interfere with the spectrum.

METHODNUJOL MULL METHOD

This method is involve straining the compound with mineral oil,nojol to create a suspension off finely ground sample in mineral oil dispersed in mineral oil. The thick suspension MULL place between the salt plates then it analyses in the spectrometer.

Disadvantage: Mineral oil obscure band over shallow band that may present in the

analyzing compound. METHOD DISSLOUTION IN ORGANIC SOLVENT

It may be used with solid which dissolve in organic solvent, must commonly CCL4.In case of mineral oil some region of spectrum are it also bind with solvent. It is also observed in organic solvent (especially at 785cm-1).

Although it is possible to cancel out the solvent from the spectrum of sample by computer.

NUCLEAR MAGNETIC RESONANCE (NMR) SPECTROSCOPY

Nuclear magnetic resonance spectroscopy involves absorption of electromagnetic radiation in the radio frequency region.

Absorption of radio waves in the presence of magnetic field is accompanied by a special type of nuclear transition, and for this reason we call this type of spectroscopy nuclear magnetic resonance (NMR) spectroscopy.

The nuclei of certain atoms behave as if they are spinning chages.Any spinning charge creates a magnetic field and behaves as if it were a tiny bar magnet, Of the three most common in organic compound, only the hydrogen nucleus proton behaves in this manner.

When a proton in an organic molecule is placed in a strong magnetic field, it can align with the field or against it

In the more stable low-energy state, it is aligned with the magnetic field. (Up field).that appears toward left on chart paper...

If energy is supplied in the form of radio waves of exactly the right frequency, radiation will be absorbed and the nucleus will flip and align against the applied magnetic field in the less stable higher energy state(downfield) .that appear right on chart paper.

The chemical shift and shielding (POSITION OF SIGNALS)

Chemical Shift (Position of Signals) The utility of NMR is that all protons do not show resonance at same frequency because, it is surrounded by particular no. of valence electrons which vary from atom to atom so, they exist in slightly different electronic environment from one another.

Position of signals in spectrum help us to know nature of protons i.e. aromatic,

aliphatic, acetylinic, vinylic, adjacent to electron releasing or withdrawing group. . When the sample absorb the energy ,however reorientation of the nucleus spin

induces a RF signal in the plane of the detector coil, and the instrument respond by recording this as a resonance signal or peak. As the field is increases linearly, the pen travel across the recording chart, AQ pen travel left to right.

A magnetic field is increases. Highly shield protons appear right of this chart, (UP

FIELD). And deshielded appear to the left. (DOWN FIELD) This is continuous spectrum instrument because in which magnetic field in the continuous fashion up field and downfield.

Spin-Spin Splitting in NMR Spectrum Peaks are often split into multiple peaks due to magnetic interactions between non-

equivalent protons on adjacent carbons; the process is called Spin-spin Splitting/Coupling. Spin-spin splitting: It occurs only between nonequivalent protons on the same carbon or

adjacent carbons. Peak: The units into which an NMR signal is split; doublet, triplet, quartet, multiplet, etc. Signal splitting: Splitting of an NMR signal into a set of peaks by the influence of

neighboring nonequivalent hydrogen. Signal coupling: An interaction in which the nuclear spins of adjacent atoms influence

each other and lead to the splitting of NMR signals.

Equivalent Protons These equivalent protons do not split each other. Protons bonded to the same carbon will split each other only if they are

not equivalent. Equivalent protons have the same chemical shift. Equivalent protons have same chemical shift without splitting occurs.

Non-equivalent Protons

If Ha and Hb are not equivalent, the splitting is observed. Nonequivalent protons on adjacent carbons have magnetic fields that may align with

or oppose the external field. This magnetic coupling causes the proton to absorb slightly downfield when the

external field is reinforced and slightly up field when the external field is opposed.

All possibilities exist, so signal is split. Non equivalent protons have different chemical shift with splitting occurs Bonds. 43Splitting is not generally observed between protons separated by more

than three bonds

Non equivalent protons have different chemical shift with splitting occur. Splitting is not observed between protons separated by more than three σ

– bonds .

INSTRUMENTATION CONTINOUSE-WAVE SPECTROMETER.

It is the basic elements of a classical 60-MHz NMR spectrometer. The sample is dissolved in a solvent containing no interfering protons (usually CCl4, CDCl3), and a small amount of TMS is added to serve as an internal reference.

The sample cell is in small is a small cylindrical glass tube that suspended in the gap between

the faces of the pole pieces of the magnet, The sample is spin around its axis to ensure that all parts of the solution experience a relatively uniform magnetic field.

Also in the magnet gap is a coil attached to 60-MHz radiofrequency generator. This coil

supplies the electromagnetic energy used to change the spin orientation of the proton .perpendicular to the RF oscillator coil is detector coil, When no absorption of energy is take place ,the detector coil pick up non of the energy given off by the RF oscillatory coil .

When the sample absorb the energy ,however reorientation of the nucleus spin induces a RF signal in the plane of the detector coil, and the instrument respond by recording this as a resonance signal or peak. As the field is increases linearly, the pen travel across the recording chart,AQ pen travel left to right .

A magnetic field is increases. Highly shield protons appear right of this chart,(UP FIELD). And deshielded appear to the left.(DOWN FIELD) This is continuous spectrum instrument because in which magnetic field in the continuous fashion up field and downfield.

The peak is calculated from frequency difference from TMS, It’s called frequency domain

spectrum. When peak is generating it ring with Cw spectrum... CW-type of NMR spectrometer is use that excite the nuclei of the isotope under observation

one type at a time. And H1 nuclei type of proton (phenyl, vinyl, methyl,) is excited individually and is resonance peak is observed and recorded independently. They excite one time individually, all other are not observed until all of the type have come into resonance.

PULSE FOURIER TRANSFORM (FT) INSTRUMENT FT-transform NMR instrument is modern type of instrument that excite the magnetic nuclei

simultaneously, E.g. isotope of the hydrogen nuclei resonance at the same time. The magnetic field uses a short (1-10micro-sec) 90MHz).This high power burst is called

pulse.

Then these nuclei emitted electromagnetic radiation, it is called free induced decay (FID). It is time domain frequency instrument.

SPECTRAL ANALYSISALKANE

ALKENE

SPECTROMETRY MASS SPECTROSCOPY

The mass spectrum of a compound helps to establish the structure of a new compound in several different ways:

1) It can give the exact molecular mass. by ionizing it and analyzing its mass using a

magnetic field. 2) It can give a molecular formula or it can reveal the presence of certain structural units

in a molecule.

TERMINOLGIES

M= Molecular ion, the ion that spectrometer analyze. This is the molecule comprised of lowest +, - atomic mass isotopes.

M/Z=Mass to charge ratio of an analyte ion .if we are asked to find the m/Z ratio for butane, what would we do?

BUTANE

Sometime a molecule distegrated during the process forming a fragment ion.

Since we desined (M) as the molecules with lowest mass atomic isotopes, we can identify fragment easily.

By studying many mass spectra we learn that this ratio can be used to determine the carbon count of analyte.

Exampe: if the relative abundance of the m+1 peak is 5.3% compared to M peak, how many carbons are in the molecule?

RA=5.3%/1.1=4.8 (5 carbon are in the analyte molecule),

M+2 RULE

NITRGEN RULE

When m/z is M is even number in formula. When m/z Is M is odd number in formula. HYDROGEN RULE

For a molecule with “C” and “N” the max number of monovalent atom (hydrogen + halogen) in the formula.

2C+N+2 DBE= double bond equivalent=C-H/2+N/2+1

INTERPRETATION OF MASS SPECTRUM

• 79Br=50.695• 81Br=49.31%

ELEMENTS AND THEIR ISOTOPES WITH THEIR RELATIVE ABUDANCE

MS INSTRUMENTATION

Each kind of ion has a particular ratio of mass to charge, i.e. m/e ratio(value). For most ions, the charge is one and thus, m/e ratio is simply the molecular mass of the ion.

SAMPLE VAPORIZATION

The basic aspect of organic mass spectrometry consist of bombarding the vapour of an organic compound with a beam of energetic electron accelerated from a filament to an energy of 70 eV to form positively charged ions (molecular ions).The heating coil is steam the sample and vaporize it.

IONIZATION

The molecules are ionised and broken up into many fragments, some of which are positive ions. In this technique, molecules are bombarded with a beam of energetic electrons.

There is electrode are place, that are positive, negative and double negative charged. Same charge repel to each other where different charge attract to each other. In this way regulate the movement of ions.

Mass spectroscopy deals with the examination of the characteristics fragments (ions) arising from the breakdown of organic molecules.

The deflection of ions, however, depends on its mass, charge and velocity.

The various positive ions, thus formed, can be accelerated and deflected by magnetic or electric fields. This fragmentation may result in the formation of an even- electron ion and radical.

DEFLACTION

The deflection is less for a heavy particle as compared to that of a light one. -The more the ion is charged, the more it gets deflected.

Though organic mass spectrometry is routinely used along with IR, NMR and

UV for structure determination, its basic theory is different from the others.

DETECTION OF IONS

When an ion hits the metal box, its charge is neutralized by an electron jumping from the metal on to the ion the beam of ions passing through the machine is detected electrically. Only ion stream B makes it right through the machine to the ion detector.

The other ions collide with the walls where they will pick up electrons and be neutralized. They get removed from the mass spectrometer by the vacuum pump.

APMLIFIER

A flow of electrons in the wire is detected as an electric current which can be amplified and recorded. The more ions arriving, the greater the current.

SPECTRUM

A mass spectrum is the plot of relative abundance of ions against their mass/charge ratio.