chroma Flashcards by Kim Grace

are multistage separation
methods in which the components of
a sample are distributed between
two phases, of which one is stationary
and the other is mobile.

Chromatographic separation
techniques

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separate different components present in a sample since samples contain a mixture
polar and nonpolar, ionic or non ionic compounds

Chromatographic separation
techniques

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may be a solid or a liquid
supported on a solid or a gel

STATIONARY PHASE

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may be packed in a column,
spread as a layer, distributed as
a film, or applied by other
techniques

STATIONARY PHASE

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may be in a gaseous or liquid
form, or a supercritical fluid

MOBILE PHASE

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any substance at a temperature
and pressure above its critical point, where distinct
liquid and gas phases do not exist, but below the
pressure required to compress it into a solid.

supercritical fluid -

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supercritical fluid -

Border of being a liquid
and being a gas.

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SP of Ion exchange

negatively charged
beads

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Separated by size by
virtue of density

Size Exclusion

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pressure needed when the column is very
packed

high pressure

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T/F: Column is a very porous material but need
help of pressure when it is very packed

TRUE

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Gas chrom SP:

liquid or solid

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Liquid Chromatography SP:

solid or liquid

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Gas chrom MP:

gas

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Liquid Chromatography MP

liquid

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a graphical representation of the
detector response,
concentration of analyte in the
eluent, or other quantity used as
a measure of eluent
concentration versus eluent
volume or time

CHROMATOGRAM

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volume between the point at
which the eluents meet and the
top of the column

DWELL VOLUME/GRADIENT DELAY VOLUME

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T/F:dwell volume: mobile phase

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Air can cause a __________ on the chromatogram

peak

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the time required for elution of an unretained component

HOLD-UP TIME (TM)

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shown as an
air or unretained solvent peak, with the baseline scale in minutes

HOLD-UP TIME (TM)

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the volume of mobile phase required for elution of an unretained
component

HOLD-UP VOLUME (VM)

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time for the unretained component to be eluted

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Volume required to push the unretained component =

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Faster flow rate - ________ components are removed

more components are removed

formula for Vm

Vm = Tm (hold-up time) x F

a measure of column efficiency

NUMBER OF THEORETICAL PLATES (N)

depends upon the substance being chromatographed as well as the operating conditions, such as the flow rate and temperature of the mobile phase or carrier gas, the quality of the packing, the uniformity of the packing within the column, and, for capillary columns, the thickness of the stationary phase film and the internal diameter and length of the column

NUMBER OF THEORETICAL PLATES (N)

Bell curve

Gaussian peaks

Used in industry

w/ electronic integrators

the portion of the chromatographic recording of the detector response when a single component is eluted from the column

PEAK

if separation is incomplete, two or more components may be eluted as one unresolved peak

PEAK

employed as a system suitability criterion in a test for related substances when baseline separation between two peaks is not achieved

PEAK-TO-VALLEY RATIO (p/v)

ratio of the adjusted retention time of a component relative to that of another used as a reference, obtained under identical conditions

RELATIVE RETENTION (r)

“unadjusted relative retention”

RELATIVE RETENTION TIME (RRT)

the separation of two components in a mixture

RESOLUTION (RS)

the time elapsed between the injection of the sample and the appearance of the maximum peak response of the eluted sample zone

RETENTION TIME (tR

may be used as a parameter for identification

RETENTION TIME (tR)

characteristic of the compounds they represent but are not unique

RETENTION TIME (tR)

Not used to characterize compounds since it is not unique but it is a parameter for identification. L

RETENTION TIME (tR)

volume of mobile phase required for elution of a component

RETENTION VOLUME (VR)

relative retention calculated for two adjacent peaks (by convention, the value of the separation factor is always >1)

SEPARATION FACTOR (⍺)

“tailing factor”, of a peak

SYMMETRY FACTOR (AS

system suitability tests is done at the

start

integral part of GC and LC methods

SYSTEM SUITABILITY TESTS

used to verify that the chromatographic system is adequate for the intended analysis

SYSTEM SUITABILITY TESTS

pH of the mobile phase (HPLC):

based on the concept that the equipment, electronics, analytical operations, and samples analyzed constitute an integral system that can be evaluated as such

SYSTEM SUITABILITY TESTS

. concentration of salts in buffer (HPLC)

ratio of components in mobile phase (HPLC): amount of the minor component (≤50%) can be adjusted ________ but cannot exceed ______- in relation to the total mobile phase

± 30%; ± 10%

wavelength of UV-Vis detector (HPLC):

± 3 nm

stationary phase column length (GC):

stationary phase column inner diameter (GC):

: ± 50%

. particle size (HPLC): -

-25% to 50% of the prescribes column length and particle size ratio

flow rate (GC):

± 50%

flow rate (HPLC):

± 50% (isocratic)

column temperature (HPLC):

± 10°

oven temperature (GC):

± 10%

oven temperature program (GC):

± 20%

A gaseous mobile phase flows under pressure through a ___________ either coated with a liquid stationary phase or packed with liquid stationary phase coated onto a solid support

heated tube

Principle of Gas Chrom: The analyte is loaded onto the head of the column via a ______ , where it evaporates. It then condenses at the head of the column, which is at a lower temperature.

heated injection port

liquid stationary phase is deposited on a finely divided, inert solid support, such as diatomaceous earth, porous polymer, or graphitized carbon, which is packed into a column that is typically 2–4 mm in internal diameter and 1–3 m in length

PACKED COLUMN

T/F: Packed Column causes less problem in sample introduction

T/F: Packed Column do not produce high-resolution chromatography

contain no packed solid support, the liquid stationary phase is deposited on the inner surface of the column and may be chemically bonded to it

CAPILLARY COLUMN

higher efficiency larger sample volume neede most commonly used carrier gas in capillary GC is helium

CAPILLARY COLUMN

A liquid mobile phase is pumped under pressure through a stainless steel column containing particles of stationary phase with a diameter of 3–10 mm (1.7 mm in ultra- high-performance liquid chromatography (UPLC))

HPLC

The analyte is loaded onto the head of the column via a loop valve and separation of a mixture occurs according to the relative lengths of time spent by its components in the stationary phase.

HPLC

most commonly used stationary phases are modified silica or polymeric beads

HPLC

beads are modified by the addition of

ong-chain hydrocarbons

(HPLC) Mobile phase in Normal Phase

(HPLC) Mobile phase in Reverse Phase

Polar

includes stainless steel, lined stainless steel, and polymeric columns, packed with a stationary phase

CHROMATOGRAPHIC COLUMN

the chromatographic technique which has seen the most intensive development in recent years, leading to improved columns, detectors and software control.

HPLC

(HPLC) The variety of columns and detectors means that the selectivity of the method can be .

readily adjusted

less risk of sample degradation Compared to gas chromatography (GC

HPLC

chroma Flashcards

(76 cards)