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BC3001: Structural & Experimental Biochemistry > Liquid Chromatography > Flashcards

Liquid Chromatography Flashcards

1
Q

Who first discovered Chromatography?

A

M. Tswett
* using CaCO3 and ethanol

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2
Q

What are the TWO main phases of Chromatography?

A
  1. Normal Phase
  2. Reverse Phase
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3
Q

What is the main principle of Chromatography?

A

All forms rely on analyte partitioning between the MOBILE and STATIONARY phases

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4
Q

What is the equation for the Distribution Coefficient?

A

Kd = [C]s / [C]m

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5
Q

What is the equation for the Partition Coefficient?

A

K = ms / mm

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6
Q

What does Chromatography rely on?

A

Differences in K for different molecules

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7
Q

What does Chromatography measure?

A

Compounds retention time

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8
Q

What is the equation for Retention Time?

A
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9
Q

What is the equation for Resolution?

A
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10
Q

What are the THREE Separation Performance parameters?

A
  1. SELECTIVITY - ability to discriminate related compounds
  2. EFFICIENCY - a measure of diffusion effects
  3. CAPACITY - amount of material that can be resolved
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11
Q

What does Low Flow Rate indicate?

A

Increased diffusion

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12
Q

What does High Flow Rate indicate?

A

Non-equilibrium partitioning

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13
Q

What is the equation for Resolution and the parameters?

A
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14
Q

What is the Isocratic elution mode?

A

Same eluent

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15
Q

What is the Gradient elution mode?

A
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16
Q

What phase is associated with Dipole-Dipole interactions during Separation / Retention?

A

Normal phase
* Al2O3, SiO2
* Apolar

17
Q

What phase is associated with Hydrophobic interactions during Separation / Retention?

A

Reverse phase
* Med polar to polar

18
Q

What is associated with Ionic interactions during Separation / Retention?

A

Ionic exchange
* High polarity

19
Q

What size beads are used in LC?

A

200-50 μm

20
Q

What size beads are used in FPLC?

21
Q

What size beads are used in HPLC?

22
Q

Put the following in order of lowest to highest pressure used
FPLC
HPLC
LC

A

LC, FPLC, HPLC

23
Q

What separation media is used in Normal Phase?

A
  • SiO2
  • Al2O3
24
Q

What separation media is used in Bonded Phase?

A

C18
*Octadecyl Silyl

25
Q

What FIVE components make up a HPLC set up?

A
  1. Solvent
  2. Pump
  3. Injector
  4. Column
  5. Detector
26
Q

What is the most common detection option in Chromatography?

A

Photometric UV-Vis

27
Q

What detection option is used for non-absorbing molecules in Chromatography?

A

Refractometric (RI)

28
Q

What detection option is used for proteins in Chromatography?

A

Fluorescent

29
Q

What column is used for peptides /oligos?

A

C18 or C8 RP

30
Q

What buffer and pH is used for Ion-Pairing Mobile Phase of peptides /oligos?

A
  • TEAA buffer
  • pH 6
31
Q

What gradient is used for peptides /oligos?

A

Acetonitrile 0 - 100%

32
Q

What detection is used for peptides /oligos?

A

UV at 260 nm

BC3001: Structural & Experimental Biochemistry (13 decks)

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