CHEMISTRY

OF

SEPARATION

OUTLINE
Introduction Types

Extraction Phase changes Electric Fields Flotation Membranes Other Chromatographic

ANALYTICAL PROCESS

Food Sampling Sample Pretreatment Extraction Calibration Instrumentation Measurement Processing Interpretation

Separation

Clean up

Concentration

Derivatization

HOMEWORK

Using the flow diagram for the analytical process, fit your research project into an analytical process.

Food, biological tissue or fluid Sampling Extraction what is the analyte? Instrumentation what will you use to measure the analyte? How will you calibrate? Processing and interpretation

INTRODUCTION
Separation

Anderson, 1987 physical transfer of a particular chemical substance from one phase or medium to another, or the actual physical separation of the components of a mixture into separate fractions. Meloan, 1999 is a process whereby compounds of interest are removed from the other compounds in the sample that may react similarly and interfere with a quantitative determination. Seader and Henley, 1998 The separation of chemical mixtures into their constituents. Separations including enrichment, concentration, purification, refining, and isolation.

INTRODUCTION

Separation Extraction Analysis

PHASE
Volatilization

Conversion of all or part of a solid or liquid into a gas What are ways that support this conversion?
Heat Strong acids Oxidation Reduction

Gas Chromtography

What analytical instrument uses this same principle?

PHASE

Distillation

The production of a vapor from liquid by heating, condensing the vapor, and collecting in a separate vessel

Vapor pressure the pressure exerted by molecules that have escaped the liquids surface Molecules in the gas state are in constant motion

Usually several hundred miles per hour This relates to surface tension

Size, shape, and chemical properties

Examples: simple, fractional

FRACTIONAL DISTILLATION

Toluene + Benzene

FRACTIONAL DISTILLATION
Plates

Theoretical plates

Represent each equilibrium step in the refluxing system

HETP (Height Equivalent to a Theoretical Plate)

Takes into account the distance from surface of liquid to the top of the column Measures the efficiency of distillation

FRACTIONAL DISTILLATION
Continuous

Refluxing

Total Partial

HOMEWORK

Ethyl isobutyrate (b.p. = 111C) and ethyl isovalerate (b.p. = 135C) are used for flavors and essences.
Briefly explain how fractional distillation works? Can these be separated using this technique?

Explain you answer? Think about theoretical plates?

AZEOTROPIC & EXTRACTIVE DISTILLATIONS

Azeotrope

Liquid mixture characterized by a maximum or minimum boiling pt. (bp) which is lower or higher than bp for any of the components and that distills without change in composition Distillation form an azeotrope

AZEOTROPIC & EXTRACTIVE DISTILLATIONS

From: Meloan, 1999. Chemical Separations: Principles, Techniques, and Experiments, John Wiley & Sons, Inc., New York.

AZEOTROPIC & EXTRACTIVE DISTILLATIONS

Extractive

A third component is added to extract one of the major components

Solvent

Other interactions

Hydrogen, dipole-dipole, ion-dipole, pi bonds

STEAM & VACUUM DISTILLATIONS

Used

for components that decompose at or near

its bp Steam

Limited to those components that are immiscible with water

Problem Emulsion form

Usually forms when densities of 2 liquids are similar Breaking emulsions Glass wool Centrifuge Salts Acids Phase separation paper (Whatman PS-1)

STEAM & VACUUM DISTILLATIONS

Vacuum

Any distillation below atmospheric pressure Advantage boiling pt differences increase at reduced pressures

SUBLIMATION
Process

which converts a solid to a gas bypassing the liquid phase A solid will sublime if its vapor pressure reaches atmospheric pressure below its melting point

SUBLIMATION
Lyophilization

ELECTRICAL FIELD SEPARATIONS

Gel Matrix

Electrophoresis
Disc Isoelectric Focusing Immuno

Capillary Electrophoresis

ELECTRICAL FIELD SEPARATIONS

Electrophoresis

Charged molecules in solution are separated based on differences in size and charge when a high voltage is applied

ELECTRICAL FIELD SEPARATIONS

Electrophoresis Theory Mobility (U) requires a net electrostatic charge Can neutral particles be separated electrophoretically? Charging processes: acids and bases, dissociation into ions by polar solvents, hydrogen bonding, chemical reactions, polarization, ion pair formation
Fs=6prhu Fs + F +

r, radius of the particle (cm), h, viscosity of the medium (poises), u, electrophoretic velocity (cm/sec)

F=QE
Q, charge on the particle E, field strength

ELECTRICAL FIELD SEPARATIONS

Electrophoresis Theory
Fs + F +

r, radius of the particle (cm), h, viscosity of the medium (poises), u, electrophoretic velocity (cm/sec)

Fs=6prhu

F=QE
Q, charge on the particle E, field strength

Thus, Fs=QE=6prhu and U=Q/6prh

ELECTRICAL FIELD SEPARATIONS

Electrophoresis
Major problem

R I W= =V I2 /R
R, V, voltage, current W,resistance, watts,R, resistance, I, I, current

Heating

Smiling

An increased rate of diffusion of sample and buffer ions leading to broadening of the separated samples. The formation of convection currents, which leads to mixing of separated samples. Thermal instability of samples that are rather sensitive to heat. This may include denaturation of proteins or loss of activity of enzymes. A decrease of buffer viscosity, and hence a reduction in the resistance of the medium.

http://www.mnstate.edu/marasing/CHEM480/Handouts/Chapters/Capillary%20Electrophoresis.pdf

FLOTATION
Purge and Trap Foam fractionation

Gas-solid flotation Liquid-solid flotation

FLOTATION
Foam

fractionation

Based on transferring one or more components in a liquid to the surface of gas bubbles passing through it and collecting the separated components in a foam at the top of the liquid.

FLOTATION

Foam fractionation

Factors
Foamers use material of opposite charge to the sample to make a good foam Defoamers benzene, quanternary amines, silicones Chain Length chain length of nonpolar end of surfactant increases, its absorption and separation increases Surfactant concentration separation increases as concentration increases up to a point pH alters ionic species

FLOTATION
Foam

fractionation

Purge and Trap

Removal and collection of volatile compounds from a liquid by diffusion of the volatiles into a stream of gas bubbles passing through it and trapping the expelled particles. Purpose - concentration

FLOTATION

Foam fractionation

Purge and Trap

Trapping System

Purging system

FLOTATION

Foam fractionation

Purge and Trap

Purge Efficiency Vapor pressure higher vapor pressure, higher purge efficiency Solubility greater solubility in the sample matrix, harder to remove Temperature increase in temperature always increases purge efficiency Sample size increase sample size requires increase in purge volume Purge volume increase in purge volume improves efficiency Purge method given same purge volume, fine bubble dispersion better than large bubbles

FLOTATION

Foam fractionation

Purge and Trap

Traps Factors for a good trap 1. Retain analytes of interest 2. Allow gases to pass readily 3. Release analyte easily 4. Stability dont release volatiles or cause side reactions 5. Reasonably priced

HOMEWORK

Explain the technique of purge and trap?

Include in your explanation

What is meant by purging and trapping? What factors influence purge efficiency? What factors influence trap efficiency?

MEMBRANES

Filtering and Sieving

Selectively remove a portion of a mixture by passing through a semi-porous material

Material if porous with small pore holes filtering Material is a screen with large pore holes screening

There is a slew of filtering papers for the analytical chemist to use

Filters with phases bonded which allows the filter to behave like a column in HPLC or GLC

MEMBRANES

Filtering and Sieving

MEMBRANES

Filtering and Sieving

Proper filtering
1. Use proper grade filter; 2. Decant; 3. Use long stem funnel; 4. Use narrow diameter stem rather than long one; 5. Use fluted funnel if possible; 6. Fold paper with 1/8 to 1/4th inch offset; 7. Tear paper at top of fold to prevent air intake; 8. Keep stem full of solution; 9. Touch end of stem to side of beaker

MEMBRANES

Osmosis & Reverse Osmosis

2nd Law of Thermodynamics systems tend toward disorder High concentration goes to low concentration Osmosis involves solvent Dialysis involves solute

MEMBRANES

Osmosis & Reverse Osmosis

Difference in thermodynamic potential Gibbs Free energy

Higher in pure solvent than solution Tendency for system to reach equilibrium free energy equal the difference is the driving force and therefore osmosis.

MEMBRANES

Osmosis & Reverse Osmosis

Application of pressure to force the solvent back to the other side Reverse osmosis Diffusion coefficient D; permeability coefficient P; solubility constant S; filtration coefficient Lp; solute permeability coefficient ; reflection coefficient

Parameters

MEMBRANES

Dialysis

Removal of low molecular weight solute molecules from a solution by passing through a semi-permeable membrane driven by a concentration gradient

Ultrafiltration
Combination of

reverse osmosis and dialysis?

OTHER TECHNIQUES

Density

Use density gradients Principle object placed in a fluid will sink if density is greater than the fluid, will float if density less than fluid or will stay suspended if densities of object and fluid are the same.
Separates based on density and amplified by applying a rotational force RCF = 1.118 x 10-5 r N2 where r, radial distance of a particle from axis of rotation in cm; N, speed of rotation in rpm

Centrifugation

HOMEWORK

Why is it not appropriate when describing centrifugation protocols to list the conditions of centrifugation in rpms?

SOLUBILITY

Extraction

Solvent

Chromatography