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CLIN CHEM LEC > CHROMATOGRAPHY, MASS SPECTROPHOTOMETRY AND LUMINESCENCE > Flashcards

CHROMATOGRAPHY, MASS SPECTROPHOTOMETRY AND LUMINESCENCE Flashcards

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1
Q

is an analytical technique commonly used for
separating a mixture of chemical substances into its
individual components, so that the individual
components can be thoroughly analyzed

A

Chromatography

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2
Q

→ could be Gas or liquid
→ Solvent moving through the column
→ Carries the sample

A

Mobile phase or carrier

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3
Q

→ Solid or liquid
→ Where the mobile phase flows
→ it does not move and stay fixed inside the column

A

Stationary phase or adsorbent

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4
Q

→ holds the stationary phase
→ it is where the sample moves

A

Column

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5
Q

→ separated components

A

● Eluate

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6
Q

→ the process of washing out a compound through a
column using a suitable solvent

A

● Elution

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7
Q

→ Mixture whose individual components have to be
separated and analyzed.

A

● Analyte

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8
Q

The time it takes for a compound or analyte to elute

A

Retention time or factor

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9
Q

● Chromatographic techniques may be classified
according to their mobile phase:

A

→ Gas Chromatography
→ Liquid Chromatography

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10
Q

separating compounds based primarily on their
volatility.

A

Gas Chromatography

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11
Q

easily evaporated at normal temperatures
● is useful for compounds that are naturally volatile or
can be easily converted into a volatile form

A

● Volatility

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12
Q

based on partition

A

Gas-Liquid Chromatography:

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13
Q

based on adsorption

A

Gas-Solid Chromatography:

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14
Q

is the emission of light by a substance

A

Luminescence

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15
Q

Give me the three types of luminescence

A

Fluorescence
Phosphorescence
Chemiluniscence

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16
Q

a beam of light is incident on certain substances, they emit visible light or radiation

A

fluorescence

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17
Q

atom or molecule that fluoresces

A

fluorophore

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18
Q

quick disappearance of fluorescence

A

quenching

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19
Q

give me the 5 basic components of fluorometry

A

light source
excitation
cuvette
emission
photodetector

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20
Q

delayed fluorescence

A

phosophorescence

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21
Q

when light radiation is incident on a certain substances they emit light continuously even after the incident light is cut off

A

phosphorescence

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22
Q

transition form a singlet to triplet state

A

intersystem crossing

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23
Q

opposite spin direction

A

Single state

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24
Q

same spin direction

A

triplet state

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25
Q

is a production of light from a chemical reaction

A

chemiluminescence

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26
Q

→ Packed columns or capillary columns

A

columns

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27
Q

very packed beads column, porous layer or
conventional inside the hollow portion of the
column.
It can be used on Gas-liquid and Gas-Solid
Chromatography

A

packed columns

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28
Q

coating (the red and blue) on the side of the
column, no particles have been added.
▪ Can only be used when using Gas-liquid
chromatography.

A

capillary or open column

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29
Q

two types of detector

A

thermal conductivity detector
Flame ionization detector

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30
Q

contain wires (filaments) that change electrical
resistance with change in temperature.

A

Thermal Conductivity detector

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31
Q

measures the unknown resistance values on
TCD.
▪ used for calibration on different instruments.

A

wheatstone bridge

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32
Q

carrier gas

A

helium

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33
Q

→ more sensitive than TC detectors.
→ uses small hydrogen flame (needed) and needs to
have a collector electrode.

A

Flame ionization detector

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34
Q

has mobile phase: (Carrier gas)

A

helium
argon
nitrogen
HAN

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35
Q

to measure analyte in a gas stream attached the
GC system
▪ universally employed where the flame is
commonly generated with hydrogen and air.

A

Flame Ionization Detector

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36
Q

introduce a sample into the inlets.
▪ manual insertion is possible.

A

Auto-Sampler

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37
Q

attached to the column head and provide the
means to introduce a sample into a continuous
flow of carrier gas

A

common inlet or injector

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38
Q

less interaction, travels faster and diluted
out first.

A

Components with less affinity of stationary
phase

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39
Q

more interaction, travels slower and diluted
later.

A

Components with more affinity to the stationary
phase

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40
Q

based on detention of ions formed during
combustion of organic compounds in a flame,
which is generated by hydrogen and air.

A

flame ionization detector

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41
Q

(where flame
produces)

A

Positive Electrode

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42
Q

(positioned
above the flame)

A

(positioned
above the flame)

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43
Q

Retention time

A

→ X axis

44
Q

Responses

A

→ Y axis

45
Q

Uses lower temperatures for separation achieving
better separation of thermolabile compounds (unstable
when heated and readily destroyed by heat)

A

Liquid Chromatography

46
Q

→ liquid-solid chromatography

A

Absorption

47
Q

acidic polar

A

silica gel),

48
Q

basic polar

A

alumina),

49
Q

nonpolar

A

charcoal)

50
Q

involved attractive forces like Hydrogen bonding
and Van der waals interaction

A

→ Absorption

51
Q

has Si-OH functional group also known as silanol,

A

Silica

52
Q

less interaction

A

release first

53
Q

More interaction

A

release last

54
Q

Liquid-Liquid Chromatography

A

Partition

55
Q

→ separation of substances according to their solubility
in an organic/non-polar solvent and in an
aqueous/polar solvent

A

Partition

56
Q

uses paper to facilitate

A

Paper Chromatography

57
Q

the distance moved by your solvent from its
starting point.

A

Solvent front

58
Q

→ Variation of L-S chromatography
→ A.k.a. size exclusion chromatography

A

Steric exclusion

59
Q

→ use of a resin (stationary solid phase) for covalent
attachment of anions or cations onto it

A

Ion-Exchange

60
Q

positively charge

A

cation

61
Q

negatively charge

A

anion

62
Q

→ most selective type of chromatography employed
→ utilizes the specific interaction between one kind of
solute molecule and a second molecule that is
immobilized on a stationary phase

A

Affinity

63
Q

Example that can be used as matrix:

A

− Agarose
− Polyacrylamide
− Polystyrene
− Cellulose
− Silica

64
Q

is attached to a spacer (-CH2-), where it
prevents nonspecific interaction of the ligand with
the matrix itself

A

Matrix

65
Q

NAD dependent dehydrogenase, kinase

A

AMP

66
Q

NADP dependent dehydrogenase

A

ADP

67
Q

Calmodulin binding enzymes

A

Calmodulin

68
Q

Biotin containing enzymes

A

Avidin

69
Q

Protein with histidine tag

A

Ni+2

70
Q

Purification of rRNA

A

Lysine

71
Q

● Variant of column chromatography
● A thin layer of sorbent, such as alumina, silica gel,
cellulose or cross-linked dextran, is uniformly coated on
a glass or plastic plate
● Most commonly used as a semiquantitative screening
test

A

THIN-LAYER CHROMATOGRAPHY

72
Q

Uses pressure for fast separations, controlled
temperature, in-line detectors and gradient elution
techniques
● Highest sensitivity

A

B. HIGH-PERFORMANCE LIQUID
CHROMATOGRAPHY

73
Q

Forces the mobile phase through the column at a
much greater velocity

A

Pump

74
Q

→ Long stainless steel
→ Silica gel – most common

A

Columns

75
Q

Can be used to introduce the sample into the path of
the mobile phase that carries it into the column

A

Sample injectors

76
Q

detect absorbances of visible
or UV light

A

Spectrophotometers

77
Q

used for spectral comparisons
and compound identification and purity and for drug
analysis in urine

A

Photodiode array

78
Q

measures current produced when the analyte of
interest is oxidized or reduced at some fixed
potential set between a pair of electrodes

A

Amperometric or electrochemical detector

79
Q

Used to record detector signal versus the time
mobile phase passed through the instrument,
starting from the time of sample injection

The graph is called

A

recorder - chromatogram

80
Q

based on fragmentation and ionization of molecules
using a suitable source of energy

A

MASS SPECTROMETRY

81
Q

→ method that requires a source of electrons in the form
of filament to which an electric potential is being
applied

A

Electron Ionization

82
Q

direct electrical current and radiofrequency voltages
of selected magnitudes are applied to two pairs of
metallic rods

A

Quadrupole mass spectrometer

83
Q

Three electrodes, in a ring shape and two end caps,
produce ions in the cavity until selectively ejected to
the ion detector

A

Ion-trap mass spectrometer

84
Q

link three quadrupoles in series

A

Triple quad

85
Q

used to scan across a preset m/z range and
select an ion of interest

A

Q1

86
Q

functions as a collision cell

A

Q2

87
Q

serves to analyze the product ions
generated in Q2 (full product ion scan or selected
reaction monitoring)

A

Q3

88
Q

chemiluminescence Reactions are oxidation reactions of

A

luminol, acridinium
esters and dioxetanes

89
Q

is a physical phenomenon that results from
the interaction of light with particles in solution

A

Light scattering

90
Q

Measures the amount of light scattered in a particulate
suspension at 90deg angle

A

NEPHELOMETRY

91
Q

the instrument of nephelometry is called

A

Nephelometer or Nephelometric
Turbidimeter

92
Q

Factors that affect Scattered Light

A

Particle size
● Concentration of particles
● Molecular weight particles
● Wavelength dependence

93
Q

→ wavelength of light > particle size
→ light symmetrically scattered around the particle

A

Rayleigh

94
Q

→ wavelength of light < particle size
→ light scatters backward but appears forward due to
destruction out of phase background scatter

A

Mie

95
Q

wavelength of light = particle size
→ more forward light scatter
→ antigen-antibody reactions

A

● Rayleigh Debye

96
Q

● Measures the amount of light blocked in a particulate
suspension

A

TURBIDIMETRY

97
Q

● Separating compounds based primarily on their volatility
● Is useful for compounds that are naturally volatile or can be
easily converted into a volatile form

A

A. GAS CHROMATOGRAPHY

98
Q

Gas-Liquid Chromatography:

A

based on partition

99
Q

Gas-Solid Chromatography:

A

based on adsorption

100
Q

packed columns (GLC/GSC) or capillary columns (GLC)

A

Columns

101
Q

→ monitor the eluate as it leaves the column
→ produce an electronic signal proportional to the
concentration of each separated component

A

Detectors

102
Q

used for spectral comparisons and compound
identification and purity and for drug analysis in urine

A

Photodiode array

103
Q

measures current produced when the analyte of interest
is oxidized or reduced at some fixed potential set
between a pair of electrodes

A

Amperometric or electrochemical detector

104
Q

▪ best and most widely used
▪ high reproducibility
▪ used at high pressures

A

loop injector

105
Q

Basic Components of Mass Spectrometry

A

● Sample inlet
● Ionization source
● Mass analyzer
● Ion detector

CLIN CHEM LEC (13 decks)

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