Temperature Programming

8/8/2019 Temperature Programming

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•emperature Programming

Isothermal separation.

If the oven temperature is kept constant during the separation, the process is •called isothermal GC chromatography.

One should know that the GC separation occurs inside the column where ..

the temperature is strictly controlled by the temperature of the oven. Oven

•emperature means column temperature and vise versa, the terms are used

interchangeably. The temperature ofthe column influences very strongly

the efficiency of the separation.

•olumn temperature is determined on the basis ofthe boiling points ofthe..

samples' components. I f the oven temperature is higher than the boiling

points ofthe eluants, they reside mostly in gaseous- mobile phase, elute too

fast and separation is not efficient. If the temperature is too low, compounds •reside mostly in the stationary phase, elute too slow, peaks are broad and •separation is not efficient. Usually the oven temperature is below or close tothe lowest boiling point of the sample components.

When the range of the boiling points of the mixture exceeds 100 0 C, it is •ery difficult to choose a proper temperature for the column. In such a case •ew method is introduced in which the column temperature varies during

the process of separation.

Variable oven temperature

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Temperature programming is the controlled change ofthe column

temperature during the process of analysis. This method is extremely

effective in the separation of complex mixtures with wide range of boiling

points

If during the process of separation the temperature of the column

fluctuates in a controlled manner the process is called temperature

programmed gas chromatography ( T P CG) . T P GC is some times

acknowledged as a separate analytical technique, however it is a naturaldevelopment of gas chromatography where the separation occurs at non-

isothermal conditions

Although the application of T P G C is relatively simple, the theory

of this techniques leads to complicated equations and difficult analysis. As

it is in other chromatographic techniques, the theory of the process is not

yet fully developed, and the equations in use now are empirical, derived

from experimental data

One can note that the temperature increases over the entire length of

the column, and thus special engineering care is taken to insure that the

temperature at each part of the column changes at the same rate as the rest

of the column

Advantages of T P GC

The advantages ofT P GC can be illustrated by comparison ofachromatograms eluted at isothermal and temperature programed conditions.

Sample is a mixture of components with a very broad range of boiling

points. Elution starts with an oven temperature below the lowest boiling

point of the components, and linearly increasing to the temperature

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..corresponding to the higher boiling point of the mixture.

At low oven temperature, early peaks, representing volatile components ..

with low boiling points, emerge as sharp symmetrical peaks, which depicts..

higher column efficiency. With the increase of the temperature compounds..

with higher boiling will start to evaporate and enter the mobile phase..

moving as compact bands at the end of the column...

..he main advantage ofT P GC is the possibility to accommodate the

separation of samples with wide range ofboiling points. Secondly, and .-most importantly, one achieves a shortening of the analytical time. .-.-Isothennal elution of a mixture, oven temperature 95 0 C; injector 220 0 C; .-detector temperature 230 0 C. .-Same sample mixture eluted under programmed temperature: .-

Initial value 95 0 C, Initial time 0 min, Rate 10 0 C / min .-inal value 115 0 C, Final time 0 min WIl

is demonstrated in Figure 44. .-he figure above shows isothennal (a) and temperature

..

WIlprogrammed (b) elution of a sample of heptane ( b. p. 110 0 C ), octane ( b.p.

1300C), and decane (b.p. 160 0 C); instmment HP 5890, column ..

Chromososrb W 5% DSCM.

T P GC requires a sophisticated control of the oven temperature, ..

gadget to perfonn the required temperature changes, and feed back

infonnation on temperature at each moment and every point of the controlled

volume.

Temperature programming allows the proper selection of

temperature resulting in total separation of all mixtures' components, and at

the same time sharply decreasing the analysis time.

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.

As one can notice in the figure bellow the elution time for the

last peak is significantly shorter than in isothennal elution. Shortening the

analytical time has a great economical effect.

Choosing proper values for the parameters of TPGC

Column length

In T P GC instruments, the length of the column is chosen considering the

required resolution. Packed columns are usually 2 - 3 meters long. Longercolumns are not beneficial in temperature progrannned instruments because

they requires bigger ovens and imposes a limit on the heating rate of the

column

Initial temperature

The initial temperature is chosen considering the lower boiling point of the

components. Initial temperature is nonnally lower than the lowest boiling

point of the mixture. The lower initial temperature has negligible effect upon

the resolution of the higher boilers since these solutes are essentially frozen

and thus unaffected by the early parts of the temperature program

Rate of temperature increase

The selection of the rate oftemperature increase is a compromise between

resolution and analysis speed. The heating rate provides the same function in

temperature progrannned analysis as the operating temperature does in

isothennal analysis .



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74 fI/IJAt lower rates, analysis time will be too long for the high boilers and band

ft!IIwidth deterioration will take place. At high rates, severe loss of resolution f!!Iwill occur.

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f!!IFinal temperature f!!I

The final temperature is the highest temperature which the oven reaches by f!!Ithis temperature rate. It has to be selected being closely below the boiling f!!I

.,point of the highest boiling point of the mixture. One has to note that even at

.,emperature lower than the boiling point of a compound, its molecules will be

distributed between gaseous phase and stationary phase, and will propagate .,

through the column. fI!!I/.,

Qualitative evaluations in T P GC .,.,

Identification of the peaks in T P CG is performed the same way ..s the identification of the peaks in isothermal elution. If the retention times ..f two compounds eluted at the same conditions (all parameters of the

..emperature program should be similar) are equivalent, this is accepted as a

proof that these compounds are identical.

..Retention time is defined also as the time which particular compound spends

inside the column during the separation process.

Identification of the compound by external standard at TP conditions.

I f a standard has the same retention time as one ofthe peaks ofthe T P GC

chromatogram, compound corresponding to this peak is accepted as identical

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to the standard.

One has to be aware that all parameters in the temperature program

for the elution of the sample have to be identical to the parameter of the

program for elution of the standards.

One ramp temperature program

Most popular temperature program include a linear increase of the

temperature from a lower to the higher point. This type ofprogram can alsoinclude an initial platform of constant temperature and/or one platform of

constant temperature at the final temperature. See Figure 45.

Change of the oven temperature during the process of separation

temperature program - is created by the operator of the GC instrument,

considering the properties of the sample.

To build a program one has to enter the values for the following parameters:

Initial temperature This is the temperature of the column at the beginning of

the analysis. One has to make an intelligent choice based on the boiling

points of the mixture. Initial temperature is similar or lower than the lowest

boiling point.

Initial time This parameter determines the time, during which the

thermostat has to hold the initial temperature constant. This time is

programmed in minutes. If one enters 0.5 that means that for 30 seconds the

oven temperature will be kept at the initial value. The length of this period

depends on the number of eluants with lower boiling points, and how close

those boiling points are. If one enters 0 min (from the Fig. 45-case a) or c)

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...then the temperature of the oven will start to increases at once. f!!!IIf!!!II

Rate Rate is the increase of the oven temperature in degreesf!!II

Celsius per minute. There are limits of the maximum rate increase. Somef!!IIJ

instruments tolerate increase of up to 30 0Cor 40

0C maximum.

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fI!!IIJFinal value Final value is the maximum oven temperature reached in

this separation process. fI!!IIJ

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Final time Final time is the time period during which the oven is kept at fI!!IIJ

temperature of the final value. fI!!IIJ,.,.,

Run time is the time taken to complete the separation of the sample mixture.

It is a sum of the initial time, time needed to reach final value, and the final

time. During this time the process of separation and analysis is completed.

The temperature program will automatically return the oven to the initial

temperature ready for the next run or experiment.

Total time includes the run time, and the time needed to cool the oven to the

initial value, and to achieve equilibrium. The instrument cools down for an

uncontrolled period. After this period has elapsed the conditions are adjusted

to those at the beginning of the run.

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Two ramps temperature program

More complicated mixtures can be eluted under a two ramp temperature

program. In this case the oven temperature is increased in two consecutive steps. To

create two ramps program one has to enter values for the following parameters:

First ramp:

1. Initial value - starting temperature of the oven;

2. Initial time - time during which the initial temperature is constant;

3. Rate - the increase of the column temperature;

4. Final value - maximum temperature reached with this rate;5. Final time the time period of oven temperature at final value

Second ramp:

6. Rate A - new rate of increase of column temperature;

7 Final value A - is the maximum temperature reached at this rate of increase of

oven temperature;

8 Final time A - is the time period during which the oven temperature is held at

it final value

After the final time is finished the run is competed. Uncontrolled cooling and

equilibrium times are needed to bring the oven temperature at the initial conditions,

ready to start the separation of another sample

Example: Set-points for two ramps program:

Initial value 80 0 C

Initial time 0.5 min

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78Rate 20 0 C / min. Rate A 15 0 C

Final value 140 0C. Final value 180 0 C

Final time 1min Final time 2 min

See Figure 47 for the graphical presentation of two ramps program

Three ramps temperature program

Temperature programming in modem instruments is designed to

give the technologist the opportunity to create even more sophisticated

temperature programs - with three ramps. The goal is to achieve full

separation at shortest analytical time. In this case oven temperature is

increased in three steps with different rates and different final times. To

create a three ramps program one has to enter values for the following

parameters:

First ramp:

1. Initial value - starting temperature ofthe oven;2. Initial time - time during which the initial temperature is constant;

3. Rate - rate of increase of the column temperature;

4. Final value - maximum oven temperature reached with this rate;

5. Final time - time period with oven temperature at final value.

Second ramp:

6. RateA - new rate of increase of column temperature;

7. Final value A - maximum temperature reached at this rate of increase of

oven temperature;

8. Final time A - time period during which the oven temperature is held at it=s

final value.

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Third ramp

9. Rate B- new rate of temperature increase of the oven;

10. Final value B - maximum temperature (or final temperature) achieved with

this rate;

11. Final time B - period of time at which the oven is held at temperature final

value B.

12. After the final time is completed, the run is stopped . Uncontrolled cooling

and equilibrium times are needed to bring the oven temperature at the initialconditions, ready to start the separation of another sample. The cooling is

initiated automatically and no specific command is needed.

Example: Example set-points for three ramps program:

First ramp Initial value 800 C

Initial time 0.5 min

Rate 20 0 C / min,

Final value 140 °C.

Final time 1 min

Second ramp

Rate A

Final value A 180 0 C

Final time A 2 min.

Third Ramp

RateB °c

Final value B 200 0 C

Final time B 1 min

See Figure 50.There are several additional requirements imposed to Temperature

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Programmed GC instrument which are not required for an instrument not

furnished with temperature programming. These requirements are:

Reqnirements for TP GC

1. Stationary phase

As has been observed it was a complicated business to choose a proper

liquid phase for a separation at isothermal conditions and this task is even

harder for T P GC. Liquid phase must remain a liquid even at the

maximum operating temperature, and not evaporate. the vaporization of the

liquid phase is often referred to as bleeding. Ifthe column bleeds it

produces noise. A common limit to the maximum vapor pressure is 10 -6 g

liquid per milliliter of carrier gas. Use oflow liquid loading and narrow

columns helps avoid bleeding.

Solidification ofthe liquid phase is also unadvisable. Freezing point or low

viscosity determines the minimum operating temperature of liquid phase.

2. Purity of the carrier gas.

For isothermal GC it is not very essential to use high purity inert gas. On

the contrary, Temperature Programmed GC requires high purity carrier gas.

Therefore, special oxygen traps and moister traps are very common

accessories to each of the modem GC instruments.

Separate column, detector, and the injection port thermostats.

It is very essential for a temperature programmed GC instrument to

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have separate heating systems for the column, the detector and the injection

port.

The thermostat that controls the temperature of the oven in T P GC

is a relatively complex and advanced apparatus. It includes a computer chip,

which collects the information, controls the temperature and executes the

program. There are also thermocouples, embedded in the walls of the oven,

which measure temperature in different parts of the oven, and sent this

information to the controller and to the electrical circuit to switch the heater

ON and OF. Separate electrical circuit controls the upper temperature limitof the column. There is also a heater indicator, that allows the technologist

to monitor constantly the temperature of the oven.

2. Wide boiling points

Majority of the samples ofnatural products, such as crude oil extract, plant

and animal oil extracts, food products and others, have very wide rage of

boiling point. In many cases the boiling points of the new products are even

not known. Isothermal elution of such type of samples will require low

starting temperature to accommodate volatile components. Viscose

components will need very long elution times several hours.

Temperature Programmed GC can analyze complex samples much faster.

Low initial temperature will favor the elution ofvolatile components.

Majority of the components with higher boiling points will stay frozenln

the beginning of the column. The increase of the oven temperature will

facilitate the elution of the higher boilers. As the oven temperature

increases furthelmore viscose samples' components will enter the gaseous

phase, and propagate through the column. Final oven temperature is

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3. Sample injection

Isothermal gas chromatography requires fast injection of the sample intofI!IJthe injection port. The temperature of the injection port is settled above the

boiling point of the higher boiler in the sample mixture. The reason for this t!'

is that all the sample components evaporate fast and enter the column as a t!'

compact band or as a "plug". This is critical if one wants to obtain better IJf!I

separation and sharp and distinguished peaks.

Fast injection of the sample is not critical for Temperature

Programmed gas chromatography. Volatile compounds will evaporate and

enter the column at once, but more viscose components will enter the f!!IJ

column slowly. They can even solidify at the beginning of the column, ifIJf!I

oven temperature, at this moment, is significantly lower than their boiling

temperature. With the increase of the oven temperature components with

appropriate boiling point will start to vaporize and enter the mobile phase.(I!fJ

One can summarize that the main requirements for Temperature f!!JProgrammed Gas Chromatography in the following manner: f!!

l . Separate thermostats

2. Computer-programmer

3. Low mass oven

4. Suitable liquid phase

5. Pure carrier gas

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Summary

Particle size of the solid support. Column efficiency is improved by the

use of small unifonn particles for solid support.

Flow rate. For maximum efficiency the column must be operated at the

optimum flow rate. This is expressed on the van Deemster plot. The minimum

HETP determines the optimum linear gas velocity.

Carrier gas. Carrier gas should have high molecular weight to achieve

optimal efficiency.

Liquid phase. Liquid phase should have low viscosity, low vapor pressure

with good solubility for the sample components.

Pressure. Optimal efficiency is obtained at low inlet to outlet ratio. The

majority of the gas chromatographs work at an outlet pressure of one

atmosphere.

Resolution can usually be improved by lowering the column temperature.

Column temperature is a very important parameter in the operation of a GC

instrument. It deserves special attention and will be discussed in detail in the

chapter on temperature programming. At this point one can summarize that

both the partition coefficient, and the diffusion coefficients are temperature

sensitive, therefore the entire process of separation is temperature dependent.

Decrease of the column temperature increases the resolution between the

peaks. Increase of the column temperature shortens the elution time.

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Temperature Programming