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PHAR 211 > Metabolomics > Flashcards

Metabolomics Flashcards

Yang

1
Q

Define metabolomics

A

Quantitative study of a whole set of small molecules (metabolome)

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2
Q

What is a primary metabolite?

A

Performs normal physiological functions such as cell growth

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3
Q

What is a intermediary metabolite?

A

Maintains homeostasis such as energy intermediates

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4
Q

What is the metabolome?

A

The complete set of small molecules, such as carbohydrates and
lipids, in a biological sample

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5
Q

The metabolome excludes: (3)

A
  1. Enzymes
  2. Genetic materials
  3. Structural molecules
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6
Q

The human metabolome is an analog of _____ ______

A

human genome

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7
Q

Metabolites are? (2)

A
  1. Final-result of cellular functions
  2. Quantifiable molecules with the closest link to phenotype
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8
Q

What are the techniques used in metabolomics? (4)

A
  1. LC-MS, especially HPLC-MS
  2. GC-MS
  3. NMR
  4. Capillary electrophoresis
    - CD-MECC
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9
Q

What is liquid chromatography (LC)?

A

An analytical technique used to separate or purify molecules dissolved in a solvent (mobile phase = liquid)

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10
Q

What are the 4 types of LC used for separation or purification of small molecules?

A
  1. Partition chromatography: paper chromatography and thin-layer chromatography
  2. HPLC – High Performance Liquid Chromatography
  3. UPLC (UHPLC) – Ultra Performance Liquid Chromatography
  4. HPLC and UPLC are commonly used in metabolomics
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11
Q

What is the equation for resolution? (Rs)

A

Rs = (V2-V1)/((W1+W2)/2)

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12
Q

The cutoff for Rs value so that the peaks are completely separate is?

A

Rs ≥ 1.5

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13
Q

What are the 3 major players of liquid chromatography?

A
  1. Resolution
  2. Column
  3. Flow rate
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14
Q

In the LC column, what are the effects of resin particle size? (2)

A
  1. Smaller particle size –> higher pressure (related to flow rate)
  2. Smaller particle size –> better resolution
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15
Q

Why is flowrate important in LC? (2)

A
  1. Loss of resolution and capacity at high flow rate
  2. Backpressure on column resin
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16
Q

How does HPLC work? (4)

A
  1. Separation of small molecule compounds dissolved in a solution
  2. Mixture of compounds is injected into the column
  3. Compounds are separated based on difference in partition
    coefficients between the mobile phase and stationary phase
    - Mobile phase: solvent
    - Stationary phase: column
  4. Mobile phase should be degassed to avoid air bubbles
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17
Q

In HPLC, what are the normal column sizes? (4)

A
  1. 5 cm
  2. 10 cm
  3. 15 cm
  4. 25 cm
18
Q

In HPLC, what is the normal particle size?

A

Normally 5 micron

19
Q

In HPLC, what is the usual pressure range?

A

200-400 bar

20
Q

In HPLC, what is the typical flow rate?

A

~1mL/min

21
Q

What is normal-phase HPLC? (2)

A
  1. Retention is decided by the interaction of the polar parts of the stationary phase and solute
  2. Packing material (stationary phase) must be more polar than mobile phase with respect to the sample
22
Q

Common column ligands in normal-phase HPLC are? (3)

A
  1. Silica
  2. CN
  3. NH2
23
Q

Common mobile phase solvents in normal-phase HPLC are?

A

Nonpolar solvents or solvent mixture such as hexane

24
Q

What is reverse-phase HPLC? (2)

A
  1. Retention is decided by the interaction of the nonpolar parts between the stationary phase and the solute
  2. Packing material (stationary phase) must be relatively nonpolar than mobile phase with respect to the sample
25
Q

Common column ligands in reverse-phase HPLC are?

A

Bonded hydrocarbons such as C18 and C8

26
Q

Common mobile phase solvents in reverse-phase HPLC are?

A

Polar solvents or solvent mixtures such as methanol and acetonitrile

27
Q

UPLC (aka UHPLC) can detect particle sizes of?
What is the column pressure?

A

1.7-1.8 micrometers
Column pressure up to 1030 bar

28
Q

True or False? Compared to UPLC, HPLC has increased resolution, sensitivity, and speed

A

False - UPLC > HPLC in these things

29
Q

What is the mobile phase of gas chromatography?
What is the stationary phase?

A
  1. Mobile phase = gas (helium most common, also hydrogen and nitrogen)
  2. Stationary phase = liquid (GLC, liquid coated on a solid) and solid (GSC, not widley used)
30
Q

When is gas chromatography used?
What does it require?
What does it do?

A
  1. Widely used in analytical chemistry
  2. Requires vaporization of compounds without decomposition
  3. Separation of gaseous and volatile substances
31
Q

GLC principle is?

A

separation by partition coefficient

32
Q

The stationary phase of gas chromatography capillary columns can be what substances? (5)

A
  1. Polyethylene glycol
  2. Polysiloxanes
  3. Hydrocarbons
  4. Esters
  5. Amides
33
Q

What is the diameter and length of gas chromatography capillary columns?

A

Diameter = 0.025-0.075cm
Length = 30-90m

34
Q

Compare helium, hydrogen, and nitrogen as gasses used for GC. Two each.

A
  1. Helium
    - Excellent thermal conductivity
    - Expensive
  2. Hydrogen
    - Better thermal conductivity
    - Reacts with unsaturated compounds and inflammable
  3. Nitrogen
    - Reduced sensitivity
    - Inexpensive
35
Q

What is the Human Metabolome Database (HMDB) of Canada?

A
  1. Detailed information of small molecule metabolites found in human body
    - Chemical data
    - Clinical data
    - Molecular biology/biochemistry data
  2. Contain 220,945 metabolite entries
36
Q

What 4 things does metabolomics study?

A
  1. Metabolite target analysis
  2. Metabolite profiling
  3. Metabolic fingerprinting
  4. Metabolic profiling
37
Q

What is metabolite target analysis?

A

Quantitative or semi-quantitative clinical and pharmaceutical analysis of a specific group of metabolites (for example, lipids)

38
Q

What is metabolite profiling?

A

Analysis of a large group of metabolites that is either related to a
specific metabolic pathway or a class of compounds
- More targeted than metabolite fingerprinting

39
Q

What is metabolic fingerprinting? (4)

A
  1. Providing information from spectra of total composition of
    metabolites
  2. 1H NMR metabolic fingerprinting
  3. Powerful method for discriminating between biological samples on the basis of differences in metabolism
  4. Disease diagnosis
40
Q

What is metabolic profiling? (2 - different from metabolite profiling I guess)

A
  1. How metabolites are organized into pathways
  2. Understand mechanism of diseases or drugs

PHAR 211 (10 decks)

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