TFH 2005 WS Chromatographie Abhängigkeit der Trennleistung eines chromatographischen Systems von...

TFH 2005 WS

Chromatographie

Abhängigkeit der Trennleistung eines chromatographischen Systems von Geschwindigkeit der mobilen Phase

Van Deemter

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TFH 2005 WS

Eddy-Diffusion A

Peakform kurz nach der Injektion

Peakform nach Passieren des Säulenbettes

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Diffusion B

Peakform kurz nach der Injektion

Peakform nach Passieren der Säule

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Stoffaustausch C

Veränderung der Peakform durch den Stoffaustausch-Prozess

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Open Tubular Columns

• At a constant pressure• Flow rate is proportional to cross sectional area• Flow rate is inversely proportional to the column length

length

areaflow

For open tubular column this means that we can get Increased linear flow rate and/or a longer columnDecreased Plate height, which means improved better resolution

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Gas Chromatography

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C-Term

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Mass Transport Band Spreading

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Asymmetric Bands

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Comparison of open tubular and packed columns.

• Open tube columns– Higher resolution– Shorter Analysis time– Increased sensitivity– Lower sample capacity

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2D - GC

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Modulators

Thermal modulator

Cryogenic modulator

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TFH 2005 WS

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Comprehensive GC

• Auch GC x GC oder multidimensional GC.

• 1. Trennung auf „normaler“ Säule.

2. Trennung auf kurzer microbore Säule.

• Gesamtes Effluenz der 1. Säule wird in sehr viele Fraktionen aufgeteilt und auf die 2. Säule gegeben.

• Resultierendes Chromatogramm hat zwei Zeitachsen (die Retentionszeiten der beiden Säulen)

• trapping und Reinjektion durch „Modulator“ bewirken Fraktionsfokussierung.

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Gas Chromatography

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Gas Chromatography

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Gas Chromatography

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Residual Solvents - ICH Topic Q3C

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HPLC - Gas Chromatography

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• She’s a reversed-phase gradient HPLC

This is Ursula

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HPLC

Injector port

guard column

LOAD / INJECT

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• Alltech 450 UV Detector (254nm fixed)

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inector detectorHead Space (80°C) MSD

oven heating rate40°C (6min) 10°C/min90°C (3min) 15°C/min135°C (3min) 15°C/min210°C (3min) 20°C/min250°C (10min)

capillary DB 624length 60 minternal diam. 0.25 mmfilm thickness 1.4µm

GC Conditions

Time needed for one cycle: 55 min

GC HP 6890

SAMPLER HS / OC

Multistage temperature program

HP 5973 - MSD

This is Pat

It is a GC-MS

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Plate THEORY

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HETP = h

In distillation,

Actual plates exist whhere vapour passes through a liquid phaseDuring this mixing, equilibrium between the phases is assumed

The height of a plate can often be directly measured

In a packed column or a capillary the plates can´t be observed – called theoretical plates

Plate theory

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Typical Plate Heights

• GC ~ 0.1 to 1 mm [30 m]

• HPLC ~ 0.01 mm [0,25 m]

• CZE ~ 0.001 mm [0,1 m]

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The Peak

• Ideal chromatographic peaks are Gaussian in peak shape.

• This comes directly from the Craig Model.

• We know certain facts about Gaussian peaks.

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Parameter aus Chromatogramm

Gaussform angenommen

k`= t´R / tR

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Plate theoryPartition coefficient k´

Solute fractions:

mobile phase: 1 / (1+ k´)

stationary phase: k´/ (1+ k´)

ß = rc/2df

1 = 1 / (1+ k´) + k´/ (1+ k´)

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TFH 2005 WS

•Alltech’s Heliflex® AT™-1ht GC capillary column is designed for the analysis of samples containing high boiling compounds

and high molecular weight samples. The phase ratio of a column () is equal to the radius of the column divided by twice the film thickness. A higher allows for the elution of high molecular weight samples in a reasonable time; or the elution of medium molecular weight samples at low temperatures. By keeping the temperatures low, the column bleed is also kept to a minimum.

•Phase Ratio: =r/2df•ID df

•0.25 0.25 250•0.32 0.25 320•0.53 1.20 110•0.25 0.10 625•0.32 0.10 800

K = k` x ß

k`= K / ß

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TFH 2005 WS

Heat as a separations tool.

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TFH 2005 WS

• We could potentially measure the width at half height and knowing it is a Gaussian peak derive the following.

22/1

2

55.5w

tN r

2

2

2

216

rr t

w

tN

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• Resolution = tr / wave = 0.589tr/w1/2 ave

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Factors Affecting Resolution

k`= K / ß

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Oberflächenchemie

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Cross section of a Fused Silica Column Open Tubular

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HTS

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PHASEN

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Choosing the Bonded Phase

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Choosing the Bonded Phase

R = C14 alkyl

R1 = isopropyl shielding

Diisopropyl side groups provide steric protection against low pH

Triple endcapping, shielding

PG = polar amide group

Polar amide group embedded in a long alkyl chain reduces interactiions between basic analytes and silica support