Lecture13

Question 0

Ion-exchange chromatography: ____ solid materials which carry ____ ions (lignitic coal, minerals, alginic acid). Ion-exchange = a ____ and ____ process retention based on attraction between ____ ____ and ____ ____ bound to stationary phase.

Answer 0

insoluble
exchangeable
reversible stoichiometric
solute ions charged sites

Question 1

Chromatographic methods and capillary electrophoresis: 6 types

Answer 1

1. ion-exchange chromatography
2. ion chromatography
3. size-exclusion chromatography
4. affinity chromatography
5. hydrophobic interaction chromatography
6. capillary electrophoresis (electric field)

Question 2

Cation-exchangers: contain bound ____ charged functional groups water softening to remove CaCl2 (hard water).
Anion-exchangers: contain bound ____ charged functional groups.

Answer 2

negatively

positively

Question 3

Further classification: strongly or weakly acidic or basic sulfonate groups remain ____ even in strongly ____ conditions carboxyl (-COOH) groups protonated pH. “Strongly basic” quarternary ammonium groups (-CH2NR3+) remain ____ at ____ pH values. “Weakly basic” tertiary ammonium groups (-CH2NHR2+) give anion exchangers ____ in moderately basic solution + ____ ability to bind ____.

Answer 3

ionized acidic <pH4
all cationic
deprotonated loose anions

Question 4

Ion-exchange selectivity: ion exchanger favor the binding of ions of ____ charge decreased ____ ____ and increased ____ (ability of an ion’s electron cloud to be ____ by nearby charges).
Donnan- equilibrium: ion exchanger placed in ____ solution-> [] of electrolyte outside _ inside ions with ____ charge as resin excluded.

Answer 4

higher hydrated radius polarizability deformed
electrolyte > same

Question 5

Conducting ion-exchange chromatography: 1. ____: small molecules ____ to temperature gradient in GC.

Answer 5

1. resins
2. gels
3. inorganic
   ion-exchange
   ionic pH
   analogous

Question 6

Ion-chromatography = high performance version of ion-exchange chromatography method ____ ____ chromatography. Problem: KNO3 and K2SO4 eventually ____, but eluate contains high [] of KOH -> high ____. Remedy: use ____ in which ____ replaced by H+. Result: KOH eluent which has high ____ converted into H2O, when analyte present: HNO3 or H2SO4 with high ____.

Answer 6

suppressed-ion anion/cation
eluted conductivity
suppressor cations
conductivity conductivity

Question 7

Ion-pair chromatography: use a reversed phase ____ column instead of ____ column -> separate ____/____ compounds.
Size exclusion chromatography: molecules separated according to ____(hydro-dynamic radius) fundamentally different from all other __ methods. ____ direct interaction of analytes with ____ phase. -> Extremely important when labile proteins (metalloproteins need to be separated/____)

Answer 7

HPLC ion-exchange polar/ionic
size LC
No stationary
purified

Question 8

Affinity chromatography: used to isolate ____ compound from complex mixture specific binding of one compound to ____ phase. Especially applicable in biochemistry and exploit highly specific interactions between: ____, ____, ____. Elution either by changing __ or ___ strength or by infusing S that has ____ affinity to ligand L.

Answer 8

single stationary
enzymes-substrates, antibodies-antigens, receptors-hormones
pH ionic stronger

Question 9

Hydrophobic interaction chromatography: principally used for protein ____ if ____ between hydrophobic protein and RP-stationary too ____ -> we could use ____ solvent to break interaction -> ____ protein, because of their size have ____ (water) and ____ regions (hydrophobic stationary phase). Adding high [] of salt (ammonium sulfate) to protein will cause protein to ___ ___ of solution.

Answer 9

purification interaction strong
organic denatures
hydrophilic hydrophobic
salt out

Question 10

Protein applied in _ M salt [] onto the column. Hydrophobic proteins eluted with ____ salt []. -> solubility of protein ___.
Capillary electrophoresis: tool of choice to observe two contamination; also separate whole ____ (diagnosis) cel nuclei vesicles and mitochondria. CE allows ____ resolution. Different ions have different ____ -> different ____. In CE (no ____ phase), CE routinely produces ____ theoretical plates.

Answer 10

1
decreasing         increases
cells      high
mobilities        speeds
stationary        50000-500000