FLAME PHOTOMETER

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instruments in biochemistry

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FLAME PHOTOMETER:

FLAME PHOTOMETER BY Dr.Brijesh Mukherjee

INTRODUCTION:

INTRODUCTION When a metal salt solution is burned, the metal provides a colored flame. Each metal ion gives a different colored flame. Flame tests, therefore, can be used to test for the absence or presence of a metal ion

IMPORTANT CONCEPTS:

IMPORTANT CONCEPTS Atomic Orbital Energy States of Atoms Ions Atomic Emission

ORBITAL ELECTRONS:

ORBITAL ELECTRONS Electrons of atoms reside in concentric spheres known as energy “shells” in which they orbit the nucleus of an atom. Each shell is assigned a principal quantum number, n . The value of n is an integer, 1, 2, 3, etc. This number determines the relative energy of the orbital and relates the distance from the shell to the nucleus---the lower the number, the lower the energy of the electron and the closer it is to the nucleus Electrons can be further distinguished according to their location in atomic orbital, specified regions in space that depend on their energies

ENERGY STATE OF ELECTRONS:

ENERGY STATE OF ELECTRONS A ground state refers to the lowest energy of system, in this case an atom. All electrons are in shells with the least energy. As energy is applied to the atom, an electron is promoted from its ground state (residence in an energy shell) to a higher energy shell or an excited state. The excited state has a higher n value, has more energy, and is less stable.

ATOMIC EMMISSION:

ATOMIC EMMISSION An emission spectra is obtained by adding energy to a material. The addition of energy promotes electrons of that material from the ground state to the excited state. As the electrons “fall” from the excited state to the ground state, they emit the energy they absorbed in the form of electromagnetic radiation (heat, light, etc.)

IONS:

IONS Ions are charged atoms; we use metal salts in this demonstration which contain metal cations and some non-metal anion For example, sodium chloride has neither sodium atoms nor chlorine atoms in it…rather, it is made of sodium ions Na + and chloride ions, Cl - The sodium ion is the cation and the chloride ion is the anion Ions undergo atomic emission and give characteristic colors This is a way to identify what ion might be present

DEFINITION:

DEFINITION A photoelectric flame photometer is an instrument used in inorganic chemical analysis to determine the concentration of certain metal ions among them sodium, potassium, calcium and lithium. It can also be defined as an device that uses flame emission spectrophotometer to measure the intensity and other properties of light Dictionary defines it as an instrument in which a spray of metallic salt in solution is vaporized in a very hot flame and subjected to quantitative analysis by measuring the intensities of the spectral lines of the metals present.

PRINCIPLE:

PRINCIPLE When an alkali metal salt is drawn into a non luminous flame, it will ionize, absorb energy from the flame and then emit light of a characteristic wavelength as the excited atom decay to the unexcited ground state. The intensity of emission is proportional to the concentration of the element in the solution. A photocell detects the emitted light and converts it to a voltage which can be recorded.

CASCADE OF EVENTS:

CASCADE OF EVENTS Atoms of elements---subjected to hot flame---specific quantum of thermal energy absorbed by orbital electrons---become unstable at high energy level---release energy as photons of particular wavelength---change back to ground state. The energy released in form of light spectra is characteristic for each element i.e.sodium emit energy at 589nm.

TABLE SHOWING EMISSION WAVE LENGTH OF VARIOUS METAL IONS:

TABLE SHOWING EMISSION WAVE LENGTH OF VARIOUS METAL IONS ELEMENT EMISSION WAVELENGTH(nm) FLAME COLOUR Sodium(Na) 589 yellow Potassium(K) 766 violet Barium( Ba ) 554 Lime green Calcium(Ca) 662 orange Lithium(Li) 670 Red

Slide 12:

Under constant and controlled conditions the light intensity of the characteristic wavelength produced by each of the atoms is directly proportional to the number of atoms that are emitting energy which in turn is directly proportional to the concentration of the substance of interest in the sample. Calcium is less excited in ordinary flames. In these cases, the amount of light given may not provide adequate sensitivity for analysis by flame emission technique. The sensitivity can be improved by using higher temperature flames.

COMPONENTS OF FLAME PHOTOMETER:

COMPONENTS OF FLAME PHOTOMETER

SCHEMATIC DIAGRAM OF FLAME PHOTOMETER:

SCHEMATIC DIAGRAM OF FLAME PHOTOMETER

FUNCTIONS OF EACH COMPONENT:

FUNCTIONS OF EACH COMPONENT NEBULIZER-means of transporting a homogeneous solution into the flame at a steady rate BURNER-maintain flame at a constant form and at a constant rate SIMPLE COLOUR FILTERS(interference type)-means of isolating light of the wavelength to be measured from that of extraneous emissions PHOTODETECTOR-means of measuring the intensity of radiation emitted by the flame

GAS MIXTURE FOR FLAME PHOTOMETRY:

GAS MIXTURE FOR FLAME PHOTOMETRY MIXTURE APPROXIMATE TEMP.(in C) Town gas air 1700 Butane air 1900 Propane air 1925 Acetylene air 2200 Town gas oxygen 2700 Propane oxygen 2800 Butane oxygen 2900 Acetylene nitrous oxide 2955

PROCEDURE:

PROCEDURE Start compressor and adjust compressed air pressure(0.2atm) Gradually open gas tap, ignite the gas and adjust gas flow to height of 2-3cm Check multimeter function by pulverizing D/W Pulverize the prepared standard solution( ie . with known concentration)one after the other into the flame and read the intensity of electrical signal Na( mmol /l) 0.25 0.5 0.75 1.0 2.0 - K( mmol /l) 0.156 0.313 0.625 0.938 1.25 2.5

Slide 18:

Create separate reference chart for Na and K using the values read earlier by drawing concentration values on x axis and intensity values on y axis. Make a reference standard curve. Pulverize the serum with the unknown concentration of Na and then K into the flame and read the intensity.(directly proportional to the intensity of emitted light)

POTASSIUM STANDARD GRAPH:

POTASSIUM STANDARD GRAPH

DATA INTERPRETATION:

DATA INTERPRETATION NORMAL RANGE -Sodium 135-145 m mol/L -Potassium 3.5-5.0 m mol/L Increased Potassium ( hyperkalaemia ) certain medicines excessive cell destruction large volume B.T. Decreased Potassium ( hypokalemia ) kidney disease excessive sweating vomiting diarrhea eating disorder

Slide 21:

Increased Sodium ( hypernatemia ) kidney disease decreased water intake diarrhea vomiting Decreased Sodium ( hyponatremia ) CCF burn patient kidney disease

USES OF FLAME PHOTOMETER:

USES OF FLAME PHOTOMETER To estimate sodium, potassium, calcium, lithium level in sample of serum, urine, CSF and other body fluids. Sodium is the main cation of plasma and interstitial fluid(extracellular) Potassium is the main cation in intracellular space Both ions are important in order to maintain the polarity of membranes of all living cells Lithium estimation is required in some psychiatric disorder where it is used therapeutically

INTERFERENCE:

INTERFERENCE In determining the amount of a particular element present, other elements can also affect the result. Such interference may be of 3 kinds- 1)Spectral-occurs when the emission lines of two elements cannot be resolved or arises from the background of flame itself 2)Ionic-in presence of each other, two elements each emit more light than they would separately 3)Formation of compounds of low volatility-as in case of phosphorous which halves the calcium emission

THANK YOU:

THANK YOU