Lecture 9-10

Question 0

ESI stand for? How does it work?

Answer 0

Electrospray ionization.
Liquid in metal capillary - taylor cone - fine filament of liquid - generate to smaller “+” droplet (relatively large droplet - smaller droplet - unstable droplet - gas-phase ion) inlet to mass spectrometer.

Question 1

LC definition?
LC creates a problem, what is it?
Solution?

Answer 1

Liquid chromatography
LC creates a huge volume of gas (as solvent evaporizes)
MS only works in vacuum, we need to remove the gas after LC prior to introduction into the MS-> need ‘interface’

Question 2

How does the improvement of MS work?
____ ____ ____ at ____ outlet combined with ____ flow of N2 creates a fine aerosol of ____ ____.
There is little ____.

Answer 2

Strong electric field, nebulizer, coaxial
charged particles
fragmentation

Question 3

Pneeumatically assisted electrospary interface for MS, for proteins, common to find multiply charged ions, ____, ____ and sometimes ____ or ____.

Answer 3

[M+nH]^n+
[M+nNa]^n+
[M+nNH4]^n+

Question 4

____ is the key to identify and quantify one/more components from complex mixtures.

Answer 4

Separation

Question 5

Solvent extraction: transfer of a ____ from one phase to another, to ____ or ____ desired analyte(s).

Answer 5

solute
isolate
concentrate

Question 6

Solution that use for extraction of aqueous solution with an immiscible organic solvent:

• lighter than water: ____, ____ and ____.
• heavier than water: ____, ____ and ____.

Answer 6

diethylether, toluene, hexane

chloroform, dichloromethane, carbontetrachloride

Question 7

Effect of pH on extraction
if solute is ___ or ___ -> molecular form and charge may change with pH.
____ ____more soluble in aqueous solution;
____ and ____ ones more soluble in organic solvent.

Answer 7

acid base
Charged species
Neutral hydrophobic

Question 8

Extraction with a ____ ____: separate metal ions from each other: selectively complex one ion with an ____ ____ and extract the complex into an ____ solvent.

Answer 8

metal chelator
organic ligand
organic

Question 9

Each ____ can react with many different metal ions, but selectivity can be achieved by controlling ___ -> select ___ where K ____ for one metal and ____ for others.

Answer 9

ligand pH

pH large small

Question 10

Partition chromatography: a technique to separate ____ ____ by repeated ____ by two ____ ____.

Answer 10

similar substances
extraction
immiscible liquids

Question 11

How does chromatography work?

Answer 11

Chromatography operates on same principle as extraction, but one phase held in place while other moves past it.

Question 12

What is mobile phase?
What is stationary phase?
What is elution?

Answer 12

Mobile: solvent (liquid or gas)
Stationary: column phase
Elution: process of passing liquid/gas through a chromatography column

Question 13

The idea behind chromatography?

Answer 13

Separate a mixture into its components based on partitioning(分隔，分开) of compounds.

Question 14

Types of chromatography:

Answer 14

Adsorption chromatography (Tswett)
Partition chromatography (Martin and Synge)
Ion-exchange chromatography (Adam and Holmes)
Size exclusion chromatography (Porath)
Affinity chromatography

Question 15

Method of adsorption chromatography:

Answer 15

Solute adsorbed on surface of solid particles: the stronger solute molecules adsorbed -> the slower it travels.

Question 16

Method of partition chromatography:

Answer 16

Liquid stationary phase bonded to solid surface (SiO2) solute partitions between stationary “liquid” and mobile phase (a gas in gas chrmatography).

Question 17

Method of ion-exchange chromatography:

Answer 17

Anions (-SO3-) or cations (-N(CH3)3+) covalently attached to stationary phase (“resin”), mobile phase = liquid
Mobile anions held near cations that are covalently attached to stationary phase.
Anion-exchange resin: only anions can be attracted to it.

Question 18

Method of Size exclusion chromatography:

Answer 18

Separates molecules according to their size, larger ones elute first followed by smaller ones.

Question 19

Method of affinity chromatography:

Answer 19

Most elective kind of chromatography: employs specific interactions between one kind of solute molecule and a second one that is covalently attached (immobilized) to stationary phase.

Question 20

Speed of mobile phase passing through chromatography column:

Answer 20

• Volume flow rate (volume solvent/unit time; ml/min)

- Linear flow rate (distance of solvent travelled/unit time)

Question 21

What’s chromatogram?

Answer 21

A graph showing detector response over elution time.

Question 22

What’s tr, tm and tr’?
What’s relative retention (alpha)? What’s k?
The ____ a compound retained by column (k=tr’/km), the ___ its k = ____ of column dimensions.

Answer 22

tr = tm + tr’ tr: retention time
tm: unretained solute tr’: adjusted retention time
alpha - separation factor
k - for each peak -> calculate retention (capacity) factor
longer larger independent

Question 23

Efficiency of separation: 2 factors determine how well compounds separated:

Answer 23

• the farther apart the better their separation

- the wider the peaks the poor their separation

Question 24

What’s resolution?
The ____ solute resides in column, the ____ the band/peak.
Inflection point has? The width of the band/peak?

Answer 24

Solute moving through column -> spreads into a Gaussian shape.
longer broader
steepest portion of curve
breadth

Question 25

Net transport of solute from region of ____ concentration to ____ concentration. By ____ ____ of molecules.

Answer 25

high low

random movement

Question 26

What measures the rate at which molecules move from region with high concentration to one with low concentration? ____ is proportional to diffusion coefficient.
Diffusion in liquid __ times ____ than in gases.
Macromolecules diffuse ____ times ____ than small molecules.

Answer 26

Diffusion coefficient
Flux = J = -D dc/dx unit: mol/m^2*s
10^4 slower
10-100 slower

Question 27

What’s plate height?

The ____ plate height, the ____ the bandwidth. GC ___mm, HPLC ___ um, CE ___ um.

Answer 27

H: A measure of column efficiency. H = sima^2/x (x: travel distance)
smaller  narrower
0.1-1.0
10
<1

Question 28

Ability of column to separate components is improved by ____ ____ ____ -> more “____ ____” = N

Answer 28

decreasing plate height
theoretical plates
N (dimensionless) N=L/H
L=column length
H=plate height

Question 29

Why bands spread?

Answer 29

Band of solute spreads as it travels through column and emerges at detector with std.dev.sima, each individual mechanism that contributes to broadening produces a std.dev.sima.

Question 30

Broadening outside the column: Solute _____ be applied in an ____ thin zone -> band has ____ width before it enters column.

Answer 30

cannot
infinitesimally(极小)
finite

Question 31

Plate height equation, also known as ____:
H ____ to variance of chromatographic band ____.
There are band-broadening mechanisms that are:___, ___ and ___.
A, B and C constants for a given column and stationary phase.

Answer 31

Van Deemter equation proportional H=sigma^2/x
H = A + B/ux + Cux
-A: Multiple paths (proportional to flow-rate)
-B: Longitudinal diffusion (inversely proportional to flow-rate)
-C: Equilibration time (independent of flow-rate)

Question 32

Explain A, B and C in van Deemter equation.

The ____ the linear flow, the ____ complete equilibrium; the ____ broadening.

Answer 32

A: arises from multiple effects for which theory murky. B: zone of solute travel to right, band becomes broader. C: comes from finite time required for solute to equilibrate between mobile and stationary phase.
slower more less

Question 33

What’s the mobile phase and stationary phase for GC? What’s the disadvantage?

Answer 33

Mobile: gas; Stationary: usually a nonvolatile liquid, but sometimes solid. Analyte: gas or volatile liquid. 1. Only ~20% of known compounds can be detected by gas chromatography. 2. If we have O2 in our gas, may oxidize the compound inside the mixture.

Question 34

How does gas-solid adsorption chromatography work?

Answer 34

Gas-solid adsorption chromatography: analyte absorbed directly on solid particles of stationary stage.
Column -> hot enough to provide sufficient vapour.