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Spectrometry

Forms of Emitted energy

Luminescence

Radiative process - energy is lost by emission of radiation

Collision deactivation

NON Radiative process

heat

energy is lost as

Flame Spectroscopy

Also called Flame photometry or flame emission spectroscopy

Flame photometry or flame emission spectroscopy

Flame Spectroscopy Also called as

flame

is used to excite the analyte before measuring the emitted radiation

Flame Spectroscopy

emission of light by atoms is characteristic for many metallic elements

potassium, sodium and calcium

This technique is also used for the assay of other elements or metals such as

589

766

554

622

670

Emission wavelength of

Na

K

Ba

Ca

Li

Yellow

Lilac

Green

Orange

Red

Flame color

Na

K

Ba

Ca

Li

(ground, excited) state.

In Flame spectroscopy, the metal cations in the solutions were initially in the

electricity, heat, EM radiation

When placed in the flame, the metals then (absorbed, emitted) energy as

(low, high) energy levels

When this occurred, electrons made transitions from (low, high) energy levels to

Flame Atomic Absorption Spectroscopy (AAS)

Both qualitative and quantitative

Flame Atomic Absorption Spectroscopy (AAS)

Uses a definite wavelength that reacts with the atoms Differs from FES in sensitivity

AAS

is more superior and specific

FES

When solution of metallic species is sprayed into flame, the fine droplets of metallic species is obtained.

FES

These neutral atoms are converted into excited state atoms by

FES

As the excited state is not stable these excited atoms returns into ground state, with the emission of radiation of specific wavelength.

FES

Wavelength and intensity of emitted transmission is measured.

AAS

When a solution of the metallic species is introduced into a flame, the droplets of metallic species will be obtained

AAS

These neutral atoms absorbs radiation of specific wavelength, emitted by hollow cathode lamp (HCL) is filled with the vapor of elements, which gives specific wavelength of radiation.

Fluorometry

For pharmaceuticals and biomolecules Use fluorophores absorbs light and emit fluorescent light

rhodamine, fluorescein

For pharmaceuticals and biomolecules Use fluorophores absorbs light and emit fluorescent light: (what fluorescent light )

Fluorescence spectrum

intensity vs wavelength

Raman Spectroscopy

Very useful for qualitative analysis - molecular structure analysis, complement to IR spectroscopy Based on Raman effect

Raman effect

light scattering

Raman spectroscopy

is similar to IR in that it is a vibrational spectroscopy technique.

Elastic scattering

Inelastic light

There are two radiation types:

Rayleigh

ex. Elastic scattering

Raman

ex. Inelastic light

Raman scattering or shift

is the energy difference between incident light and scattered light, this is notated as the intensity

Raman shift

provides structures present in the sample

Stokes scattering

can be observed in the lower wavenumber or longer wavelength region and the opposite for the Antistokes

Nephelometry

It is an analytical chemistry technique used to measure the amount of turbidity or cloudiness in a solution caused by the presence of suspended insoluble particles.

Nephelometry

intensity of the scattered light

Nephelometry

It is best suited and offers higher sensitivity for the analysis of small, suspended particles at low concentrations

Turbidimetry

A technique is based on the measurement of the loss of intensity of transmitted light in an emulsion (or solution containing fine particles) due to the scattering effect of particles suspended in it.

Turbidimetry

It is usually applied to relatively large insoluble particles at high concentrations

Wavelength

does not matter since no absorption takes place

Halogen and xenon lamps or lasers

can be used as light sources

Turbidimeter

For__, a detector is placed opposite to the light source and at an angle relative to the incoming light beam

Nephelometer

For __, a detector is placed at a 90 degree- angle

Nephelometry

Pharmaceutical industry, it is used to assess the solubility of drugs or compounds, for quantification of microbial growth, or cell count of microorganism

Turbidimetry

-Assay of antibiotics (Chlortetracycline, doxicycline, gentamicin), calcium panthothenate, vitamin B12, Excess of chlorides and sulfates

Colorimetry

Branch of spectrophotometry in which absorption measurement is made in the visible region of the spectrum.

Colorimetric methods

are often used even though a compound ( or its colored derivative) to be analyzed absorbs in the UV region.

Beer - Lambert's laws

Colorimetry Works on the basis of__

filter photometer

Colorimeter is also called

1. Low cost

2. Simple instrument

3. Use for qualitative and quantitative analy

Advantages of Colorimetry:

glucose oxidase

Blood glucose reacts with

catalytic action

Gluconic acid and H2O2 forms as a result of the

Nuclear Magnetic Resonance

transitions between energy levels may be generated by radiant energy if the molecules are first placed in a magnetic field.

NMR

The principle behind this is that many nuclei have spin and all nuclei are electrically charged.

Spin

is a fundamental property of nature like electrical charge or mass

Spin

is a measure of angular momentum

Nuclear MagneticResonance

If an external magnetic field is applied, an energy transfer is possible between the base energy to a higher energy level (generally a single energy gap).

NMR spectrum

is the plot of radio frequency applied against absorption

Resonance

signal in the spectrum

nuclear magnetic resonance

spectroscopy experiment involves using energy in the form of electromagnetic radiation to pump the excess alpha oriented nuclei into the beta state.

nuclear magnetic resonance

is a research technique that exploits the magnetic properties of certain atomic nuclei.

NMR analysis

is used to confirm the identity of a substance.

Chemical shift

- frequency of the signal expressed with reference to a standard

Chemical shift

Computed as the difference between the resonance frequency of the nucleus and a standard, relative to the standard.

Chemical shifts

are typically reported as a fraction of the nominal resonance frequency or parts-per-million (ppm) are used.

NMR spectroscopy

this standard is often tetramethylsilane, Si(CH3)4, abbreviated TMS.

Spin-spin coupling

is a phenomenon that takes place due to the neighboring proton (H or C).

spin-spin coupling-peaks

are split into multiplets which depend upon the groups on adjacent nuclei.

Spin spin coupling

occurs between non-equivalent hydrogens

Splitting

occurs primarily between hydrogens that are separated by three bonds

Splitting

is most noticeable with hydrogens bonded tocarbon

J coupling or coupling constant

is the difference between the adjacent sub-peaks in a split signal

INTEGRATION

HOW TALL OR BIG THE RESONANCE

INTEGRATION

TO DETERMINE HOW MANY CHEMICALLY EQUIVALENT PROTON WILL GENERATE THAT PEAK.

SPLITTING

DETERMINE HOW MANY PEAKS (SINGLET, DOUBLET, TRIPLET, QUADRUPLET)

SPLITTING

DEPENDING ON THE NEIGBORING PROTON

Mass Spectrometry

It is an instrumental method for identifying the chemical constitution of a substance by means of the separation of gaseous ions according to their differing mass and charge.

mass spectrometer

generates multiple ions from the sample under investigation, it then separates them according to their specific mass-tocharge ratio (m/z), and then records the relative abundance of each ion type.

Ion Source:

Analyzer:

Detector System

The instrument consists of three major components:

Ion Source

For producing gaseous ions from the substance being studied.

Analyzer

For resolving the ions into their characteristics mass components according to their mass-to-charge ratio.

Detector System

For detecting the ions and recording the relative abundance of each of the resolved ionic species

Acceleration

The ions are accelerated so that they all have the same kinetic energy

Deflection

The ions are then deflected by a magnetic field according to their masses. The lighter they are, the more they are deflected.

mass spectrum

measures the masses within a sample

Mass spectrometry

is used in many different fields and is applied to pure samples as well as complex mixtures.

mass spectrum

is a plot of the ion signal as a function of the mass-to-charge ratio.