Adriaan Ampe Promotor: Prof. Dr. Frederic Lynen

Supervisor: Mathijs Baert

A dissertation submitted to Ghent University in partial fulfilment of the requirements for the degree of Master of Science in Chemistry Academic year 2018-2019

Development Of Chiral And

Achiral Polymerized Ionic

Liquid Stationary Phases

For High Temperature Gas

Chromatography

i

Acknowledgements

Working on this thesis took more effort than initially assumed, mainly since it felt like every

time progress was achieved on the experiments, one or more other issues arose. There were

moments in which I just wanted to drop all the work and give up on the thesis subject. But

luckily during those harsh times when they arrived I could always depend on a variety of

people to help me out. Despite how difficult it is to properly convey my feelings of gratitude

towards all the people that helped me during this year on paper, I will still try to do so.

First and foremost, I want to thank the promotor of this work, Prof. Dr. Frederic Lynen for

allowing me to expand upon the research I assisted in during my bachelor project. Secondly, I

cannot thank you enough for the variety of new ideas and possibilities for further

experiments every time you were updated on the work.

A special thanks also goes out to the two supervisors of this thesis, Mathijs Baert and Ing.

Pieter Surmont. Mathijs, I’m not sure I would have managed to get through all the synthesis

to obtain the polymers required without your insight and ideas on how to perform certain

synthesis steps and coming up with possible ways of purification in between different

synthesis steps. Also, your help with deciphering the more complex 1H-NMR spectra proved

invaluable. Pieter, a big thanks for helping me refresh the basics of the GC-FID and helping

whenever any issues with the equipment came up.

Another big thanks to Dr. Thomas Van De Velde for helping with measurements and

optimisation for the measurements on the comprehensive GCxGC-FID.

I also want to thank my parents and my sister for being the moral support from home.

Despite the fact they barely could help me out with specific issues I had with my thesis, they

still tried their best to assist in thinking ahead whenever something went wrong.

ii

A great many thanks also to my fellow thesis students, Koen Arys, Ine Mertens, Sander

Smeets, and Phanumat Latsrisaeng with whom I shared the office with during the year who

had to bear the complaints whenever something went wrong, but who also helped to think of

possible solutions on issues with synthesis or purification.

I want to thank all my friends for helping me with temporarily taking my mind off my thesis,

which allowed me to fully relax when I needed it the most.

Finally, I also want to thank Benjamin Bazi for proof-reading my thesis and filtering out

possible mistakes in layout or phrasing.

I mean this from the bottom of my heart when I say that I wouldn’t have been able to do this

without the help of everyone mentioned above.

iii

Contents

Acknowledgements .................................................................................................................................. i

Contents .................................................................................................................................................. iii

1 Introduction ..................................................................................................................................... 1

2 Fundamental Aspects in Gas Chromatography ............................................................................... 5

2.1 Retention ................................................................................................................................. 6

2.2 Graphical resolution ................................................................................................................ 8

2.3 Column efficiency .................................................................................................................... 9

2.4 Chromatographic resolution ................................................................................................. 15

2.5 Peak capacity ......................................................................................................................... 16

2.6 Comprehensive GC (GCxGC) .................................................................................................. 17

3 Chirality and chiral separations ..................................................................................................... 19

3.1 Chirality ................................................................................................................................. 19

3.2 Thermodynamics of chiral separations ................................................................................. 22

4 Ionic liquids .................................................................................................................................... 23

4.1 Introduction to ionic liquids .................................................................................................. 23

4.2 Development of IL stationary phases .................................................................................... 25

4.3 Applications of IL stationary phases ...................................................................................... 29

5 Synthesis and application of chain-grown polymeric ionic liquids ................................................ 33

5.1 Synthesis of chain-growth monomer .................................................................................... 33

5.1.1 Monomer synthesis ....................................................................................................... 33

5.1.2 Polymer synthesis .......................................................................................................... 34

5.1.3 Ion exchange reaction ................................................................................................... 35

5.2 Coating of chain-growth ionic liquid polymer ....................................................................... 35

5.3 Results and discussion ........................................................................................................... 38

5.3.1 Assessment of thermal stability .................................................................................... 38

5.3.2 Evaluation of retention and separation ........................................................................ 41

5.3.3 Assessment of column efficiency .................................................................................. 43

5.3.4 Assessment of chiral separations .................................................................................. 45

5.3.5 Influence of thickness of coating ................................................................................... 46

5.3.6 Comprehensive GC measurements (GCxGC-FID) .......................................................... 47

iv

6 Synthesis and application of step-growth polymeric ionic liquids ................................................ 49

6.1 Synthesis of step-growth polymer ........................................................................................ 49

6.1.1 Monomer synthesis ....................................................................................................... 49

6.1.2 Polymer synthesis and ion exchange reaction .............................................................. 51

6.2 Coating of step-growth polymer ........................................................................................... 52

6.3 Results and discussion ........................................................................................................... 52

6.3.1 Assessment of thermal stability .................................................................................... 52

6.3.2 Evaluation of retention and separation ........................................................................ 53

6.3.3 Assessment of chiral separations .................................................................................. 55

7 Summary and conclusions ............................................................................................................. 57

Appendix................................................................................................................................................... I

A Reagents and materials ............................................................................................................ I

B Equipment ............................................................................................................................... II

B.1 NMR spectroscopy .............................................................................................................. II

B.2 Gas chromatography – Flame ionization detector (GC-FID) ............................................... II

B.3 Comprehensive GCxGC-FID ................................................................................................. II

C NMR-spectra ........................................................................................................................... III

C.1 Monomer CPIL2 .................................................................................................................. III

C.2 CPIL2 ................................................................................................................................... IV

C.2 Synthesis CPIL3 ................................................................................................................... IV

Bibliography ............................................................................................................................................. V

1

Chapter 1

Introduction

The field of separation science is one of significant importance in chemistry, after all most

other fields which have connections to organic synthesis frequently require the use of

separation sciences, in one way or the other, to identify and/or purify their molecules of

interest. Many methods incorporated in other branches of chemistry are part of the branch

of separation sciences in the truest sense. Organic chemists for example make use of many

methods to follow their reaction (e.g. Thin Layer Chromatography, TLC) or purify their

product by means of a column, either the old analogue version of Liquid Chromatography or

also Liquid Chromatography coupled with either a UV-detector or a mass spectrometer.

Macromolecular chemists make use of analytical separations such as SEC (Size-Exclusion

Chromatography) to determine the degree of polymerisation and dispersity’s of their

polymers. Even outside of the field of chemistry, separation sciences play a crucial role in the

separation, identification and/or quantification of various (complex) organic monsters.

Without the existence of separation sciences, routine analysis of compounds in (complex)

samples such as the blood analysis and quality control of products would be nearly

impossible.

Large industries such as the pharmaceutical industry also depend a lot on the progress made

in the field of separation techniques, it is e.g. thereby highly relevant to separate, identify

and quantify achiral and chiral impurities, next to the desired active pharmaceutical

ingredient which could have either a harmful effect (carcinogenic, toxic, …) or at best no

effect at all. Nowadays, chiral separations are usually performed by means of (high

performance) liquid chromatography (HPLC). This is in part due to the columns developed

for the latter being highly optimized for the typically semi-polar pharmaceutical solutes and

due to limited maximum operational temperatures of commercially available chiral

stationary phases for gas chromatography (~230°C for Supelco® columns1).

2

However, the use of HPLC for analysis is inherently more limited in terms of achievable

efficiency and resolution when compared to what can be reachable by gas chromatography.

The main reason therefore is the much higher amount of theoretical plates which can be

reached in GC compared to liquid driven chromatography. Next to this, separations in GC are

typically more cost effective and can also be considered greener compared to LC since no

liquids, generally organic solvents, are required for use as a mobile phase2. However, a

limitation of gas chromatography is the thermal limits of the columns used in e.g. chiral GC.

Therefore, improvement towards more efficient (chiral) GC columns capable of better

separations is relevant. Since an increase in thermal limits of (chiral) separations in GC would

lead to greener ways to analyse solutions. In the long history of separation science, the

discovery of ionic liquids and application thereof as stationary phases in GC and as stationary

and/or mobile phases in LC has allowed for significant breakthroughs regarding e.g. the

increase of thermal limits of the ensuing chromatographic methods.

These ionic liquids are considered “new types of molecules” which have shown several

interesting physical and chemical properties for the application in separation science. When

used as stationary phases in GC, a “dual” nature characteristic has been observed3: they

separate polar compounds as if they were a polar stationary phase and non-polar

compounds as if they were a non-polar stationary phase. This interesting property,

combined with their high thermal stability and the tuneability of ionic liquid stationary

phases, has made them a good candidate to potentially achieve higher-temperature

separations. The advantage of their tuneability is the fact that this kind of stationary phase

can be synthesized to be more polar when separation requires so, but it also allows for

synthesis of high temperature chiral stationary phases and are therefore a good candidate

for the improvement upon the current generation of chiral stationary phases available for

GC.

Previous reports4 have shown that ionic liquids based on alkyl-imidazolium cations can be

synthesized with thermal stabilities up to those of conventional (non-polar) PDMS-stationary

phases, which have maximum operational temperatures of up to 350°C. Yet, in comparison

to these PDMS-stationary phases, these ionic liquid stationary phases boast a better

selectivity and resolution towards polar compounds, making them more versatile.

3

In this master dissertation, attempts will be made to push the limits of (chiral) gas

chromatography by synthesis of new chiral polymerised ionic liquids which boast a higher

maximum operational temperature to potentially replace the conventionally used stationary

phases currently applied in gas chromatography.

4

5

Chapter 2

Fundamental Aspects in Gas Chromatography

Gas chromatography is the leading method for analysis of volatile organic components.

Separations are achieved on differences in boiling points and/or by interaction of analytes

with the stationary phase. A gas chromatographic system consists of three major parts: an

injector system, a column in a thermally controlled oven, and a detector (Figure 2.1):

Figure 2.1: Schematic representation of a basic GC system.

Conventionally used columns used in gas chromatography range from apolar, PDMS-

columns, to more polar, polyethylene glycol and WAX columns. The two most used detectors

are either a flame-ionization detector (FID) or a mass spectrometer. In which the mass

spectrometer is generally used for qualitative measurements whereas an FID is

conventionally applied for quantitative analysis.

6

2.1 Retention

The retention time, tr, is the amount of time a compound spends on the column after it has

been injected and is determined by the time between injection of the compound and

detection. It consists of the dead time or hold up time of the column, t0, and the net

retention time, tr’:

𝑡𝑟′ = 𝑡𝑟 − 𝑡0 (2.1.1)

The dead time is the time required for a non-retained compound to travel through the

column. In practice the elution time of the solvent peak can be used to determine the dead

time. The net retention time describes the affinity of a component for the stationary phase

and is a measure for how much this component is retarded by interaction with the

stationary phase. In practice, the net retention time is rarely used since this value always

depends on the dead time of a column. Therefor the net retention time is usually

transformed into a more absolute value by dividing it by the void time t0:

𝑘 =

𝑡𝑟′

𝑡0=

𝑡𝑟 − 𝑡0

𝑡0 (2.1.2)

Doing so yields the retention factor, which expresses how much longer a sample component

is retarded by the stationary phase compared to how long it would take for the component

to travel through the column with the velocity of the mobile phase.5 The retention time and

retention factor are influenced by several factors such as volatility of the component, the

stationary phase, amount of stationary phase, column diameter, column length, mobile

phase, mobile phase velocity and column temperature.6

7

From the retention factors of two peaks, the selectivity factor, α, can be calculated as:

𝛼 =

𝑘2

𝑘1=

𝑡𝑟2− 𝑡0

𝑡𝑟1− 𝑡0

(2.1.3)

In which k1 and k2 are respectively the first and last eluting peaks in the chromatogram. This

selectivity factor is a measure for the ability of a chromatographic system to chemically

distinguish between two specific sample components and is a measure for the distance

between the apices of both peaks. In which high α-values indicate good separation power

and hence good separation between the apices of the two peaks. However, the selectivity

factor is limited since it doesn’t allow to learn anything in how far peaks in a chromatogram

are separated (and hence resolved).7

Figure 2.2: Chromatogram showing the representation of dead time, t0, and retention time, tr.

t0

tr

8

2.2 Graphical resolution

Therefor resolution is a better measure of the quality of separation between two (closely)

eluting compounds. It can be obtained in either a graphical (practical) or chromatographic

(theoretical) (see §2.4) way. The resolution between two adjacent peaks can be obtained

from a chromatograph in a graphical way by the following formula:

𝑅𝑆 =

𝑡2 − 𝑡1

12 (𝑤𝑏1

+ 𝑤𝑏2) (2.2.1)

Where t2 and t1 are the retention times of the second and first peaks respectively and wb,1

and wb,2 are the widths of these peaks measured at the baseline.

In practice, resolution is generally only calculated for closely eluting peaks, meaning that the

widths of both peaks usually are approximately equal, which simplifies the formula for

resolution above into:

𝑅𝑆 =

𝑡2 − 𝑡1

𝑤𝑏2

(2.2.2)

As shown in Figure 2.3 below, a minimal resolution of 1.5 is required to achieve baseline

separation for two closely eluting peaks.

Figure 2.3: Graphical representation of resolution showing the effectiveness

of separation based on resolution.6

9

2.3 Column efficiency

The term column efficiency is, as the name implies, a measure to determine the efficiency of

a specific column or stationary phase.

It is reported as the number of theoretical plates or plate number, N, which is a concept

which was borrowed from fractional distillation.8 The plate number is calculated from a

peaks’ standard deviation, σ, and elution time, tr, as:

𝑁 = (

𝑡𝑅

𝜎)

2

= 16 (𝑡𝑅

𝑤𝑏)

2

= 5.54 (𝑡𝑅

𝑤ℎ)

2

(2.3.1)

The peak width at baseline, wb, and at half height, wh, are considered equal to 4σ and 2.355σ

respectively due to a perfectly eluting peak having a Gaussian shape.

Since longer columns intrinsically contain more plates than a shorter column, it’s clear that

the plate number N can only be used as a parameter if columns with equal length are

compared to each other. To overcome this limitation, another variable is generally used,

called the height equivalent to a theoretical plate (H or HETP), which describes the column

efficiency independently of the length.

𝐻 =

𝐿

𝑁 (2.3.2)

The smaller the plate heights, the greater the number of theoretical plates a column can

contain and therefor the higher the efficiency of the column.

The HETP value for a column is also dependent on the mobile phase velocity and mobile

phase itself. The influence of the mobile phase velocity on the height of a theoretical plate

has been described by Van Deemter in 19569 by means of the Van Deemter equation:

𝐻 = 𝐴 +

𝐵

𝑢+ 𝐶𝑢 = 𝐴 +

𝐵

𝑢+ (𝐶𝑀 + 𝐶𝑆)𝑢 (2.3.3)

Where A is the Eddy diffusion, B is the longitudinal (axial) diffusion and C is the resistance to

mass transfer consisting of a mobile phase and stationary phase component denoted as CM

and CS respectively.

10

The Eddy diffusion term (=turbulent flow diffusion), A, describes the diffusion in a column

caused by the different pathways a molecule can take to travel through a packed column

(Figure 2.4). This term is dependent on the particle diameter of the column packing, dp, and

the packing quality, λ:

𝐴 = 2𝜆𝑑𝑝 (2.3.4)

Figure 2.4: Illustration of the Eddy diffusion phenomenon in packed columns.10

For open tubular capillary columns, no packing is present and therefor A is equal to 0.

Longitudinal diffusion, B, is a band broadening process by diffusion due to a concentration

gradient, therefor diffusion toward and opposed to the direction of the flow of the mobile

phase. This factor is influenced by the diffusion coefficient of the mobile phase, DM, and an

obstructive factor, γ, which describes the hindering of diffusion by the column packing

material (in packed columns):

𝐵 = 2𝛾𝐷𝑀 (2.3.5)

Figure 2.5: Illustration of the longitudinal diffusion occurring in a column.11

It is important to note that for open tubular columns, the obstructive factor is equal to 1,

since no packing material is present.

11

Both resistance to mass transfer factors CS and CM are dependent on complex functions of

the retention factor k, column design parameters, and the diffusion coefficients in both

phases. More specifically, the mass transfer term for the stationary phase, CS, is dependent

on the square of the film-thickness on the supporting material df² and inversely proportional

to the diffusion coefficient in the stationary phase DS:

𝐶𝑆 =

𝑓𝑆(𝑘)𝑑𝑓2

𝐷𝑆 (2.3.6)

The mobile phase mass transfer term, CM, depends on the square of the column radius, rc,

and inversely proportional to the mobile phase diffusion coefficient DM:

𝐶𝑀 =

𝑓𝑀(𝑘)𝑟𝑐2

𝐷𝑀 (2.3.7)

The figure below represents how the mass transfer phenomenon causes band broadening. 12

Figure 2.6: Representation of the broadening caused by resistance to mass transfer.12

(a) distribution of compound, (b) movement of compound in mobile phase due to flow causing disturbed

equilibrium, (c) reestablishment of the equilibrium (d) new broader distribution of the compound.

When considering the Van Deemter equation above, the conclusion can be made that at low

mobile phase velocities the longitudinal diffusion coefficient, B, will have the largest

influence on the plate height, whereas at higher mobile phase velocities, the mass transfer

coefficients will have the most significant influence on the increase of the plate height.

12

Since a minimal plate height and therefore optimum mobile phase velocity is preferred, this

value can be calculated from the Van Deemter equation by taking the first derivative with

respect to mobile phase velocity, u:

𝐻′ = −

𝐵

𝑢2+ 𝐶 (2.3.8)

Setting the equation above equal to 0 then allows for calculation of the optimal velocity,

which is the velocity at which H reaches a minimum:

𝑢𝑜𝑝𝑡 = √𝐵

𝐶 (2.3.9)

By filling this back into the Van Deemter equation and assuming an open tubular capillary

column, which means no packing is present and therefore Eddy diffusion is absent (𝐴 = 0),

the Van Deemter equation can be written as:

𝐻 =

𝐵

𝑢+ 𝐶𝑢 =

𝐵

𝑢+ (𝐶𝑀 + 𝐶𝑆)𝑢 (2.3.10)

It can now be observed that the minimal plate height for a column is obtained as:

𝐻𝑚𝑖𝑛 = 2√𝐵𝐶 (2.3.11)

When completely working out the Van Deemter equation (Equation 2.3.10) by filling in the

equations describing the longitudinal and resistance to mass-transfer terms (Equations 2.3.5

to 2.3.7), the Golay equation is obtained as:

𝐻 =

2𝐷𝑀

𝑢+ [

1 + 6𝑘 + 11𝑘2

24(1 + 𝑘)²𝐷𝑀

] 𝑟𝑐2𝑢 + [

2𝑘

3(1 + 𝑘)2]

𝑑𝑓2

𝐷𝑆𝑢 (2.3.12)

For narrow columns coated with thin films, the resistance to mass transfer in the stationary

phase contribution can be neglected (CS = 0) and the Golay equation can be simplified to

obtain:

𝐻 =

2𝐷𝑀

𝑢+ [

1 + 6𝑘 + 11𝑘2

24(1 + 𝑘)²𝐷𝑀

] 𝑟𝑐2𝑢 (2.3.13)

13

This worked out equation shows that the plate height is dependent on the diffusion

coefficient of the analyte, the retention factor, the column diameter and on the linear

velocity. Calculating Hmin with these parameters then yields:

𝐻𝑚𝑖𝑛 = 2√𝐵𝐶 = 𝑟𝑐√1 + 6𝑘 + 11𝑘²

3(1 + 𝑘)² (2.3.14)

When considering well retained components, the following step can be performed:

lim𝑘→+∞

𝐻𝑚𝑖𝑛 = lim𝑘→+∞

𝑟𝑐 √1 + 6𝑘 + 11𝑘²

3(1 + 𝑘)²≈ 𝑟𝑐√

11

3≈ 2𝑟𝑐 = 𝑑𝑐 (2.3.15)

Which leads to the conclusion that for the large values of k, the minimal plate height, Hmin,

will be equal to the diameter of the column. By implementing the value found above in

Equation 2.3.14, the efficiency of a column can be predicted as:

𝑁 ≈

𝐿

𝑑𝑐 (2.3.16)

This simplifies the Golay equation significantly by allowing efficiency of a GC column to be

estimated based on only the length and diameter of the capillary.13

14

As expected, also the mobile phase used has an influence on the plate height of a column. In

Figure 2.7 below, a comparison is made between three conventionally used mobile phases in

gas chromatography (N2, He, and H2) by means of the Van Deemter curves they lead to.

From this figure, one can see that the minimal plate heights are comparable for all three

gases, but that the use of H2 as carrier gas is advised as it allows for the highest optimal

velocity. This allows for faster measurements and enhanced signal to noise ratios obtained in

the chromatogram.

Figure 2.7: Influence of carrier gas in the Van Deemter curve.14

15

2.4 Chromatographic resolution

In contrast to graphical resolution (§2.2), resolution can also be written down in a more

theoretical manner by substituting the retention time and peak width in the equation for

graphical resolution (§2.2; Equation 2.2.1). This by making use of the definitions of the

retention factor (§2.1; Equation 2.1.2), plate number N (§2.3; Equation 2.3.1), and of the

selectivity factor α (§2.1; Equation 2.1.3). This yields the following equation:

𝑅 =

√𝑁

4

𝛼 − 1

𝛼

𝑘2

1 + 𝑘2

(2.4.1)

The equation above (Equation 2.4.1) is also referred to as the master equation of

Chromatography. This formula has a significant importance since it shows how column

efficiency (N), selectivity efficiency (α) and retention factor influence the resolution of a

chromatogram. From this equation it can e.g. be concluded that to double the resolution, a

fourfold increase of the plate number is required. A more straight-forward way to increase

resolution would be to increase either the selectivity factor or the retention factor when

possible. However, in practice, resolution is only typically calculated for critical pairs and

optimising the separation of these critical pairs can have a negative influence on the

separation of other compounds in the chromatogram.

16

2.5 Peak capacity

Peak capacity is a measure of the capacity of a method to separate a number of peaks

shoulder-by-shoulder over the timespan covered by the analysis. For isocratic/isothermal

separations, the peak capacity of a column is given by:

𝑃𝐶 = 1 + ∫

1

4𝜎𝑑𝑡

𝑡2

𝑡1

= 1 + ∫1

4 (𝑡𝑅

√𝑁)

𝑑𝑡𝑡2

𝑡1

(2.5.1)

𝑃𝐶 = 1 +

√𝑁

4ln

𝑡2

𝑡1= 1 +

√𝑁

4ln

𝑘2 + 1

𝑘1 + 1 (2.5.2)

Where σ is the standard deviation (𝜎 =𝑡𝑅

√𝑁) of a peak, N is the plate number (§2.3) and k is

the retention factor of the peak (where k2 is the last eluting peak in the set timeframe, and

k1 is the first). In temperature- or solvent-gradient separations, this formula can be rewritten

because all peaks have approximately the same width:

𝑃𝐶 = 1 + ∫

1

�̅�𝑑𝑡

𝑡2

𝑡1

⟹ 𝑃𝐶 = 1 +𝑡2 − 𝑡1

�̅� (2.5.3)

Where W̅ is the average peak width in the chromatogram. If the time interval covers the

entire gradient range 𝑡2 − 𝑡1 can be replaced with the gradient time tg.

Therefore, the formula above can be rewritten as 15,16:

𝑃𝐶 = 1 +

𝑡𝑔

�̅�

(2.5.4)

Figure 2.8: Schematic example of a peak capacity of 40.17

17

2.6 Comprehensive GC (GCxGC)

A more recent development in gas chromatography is comprehensive gas chromatography.

The latter is one of the types of 2-dimensional (2D) chromatography whereby one or more

fractions are collected from a first-dimension separation (1D) and reanalysed in second

dimension separation (2D) depicting different selectivity. In a comprehensive GC system

(GCxGC), all fractions eluting from the 1D are reanalysed in real time in 2D by fast GC on short

columns. Important is thereby that the separation occurs over two different stationary

phases depicting complementary characteristics e.g. combination of an apolar and a polar

stationary phase.

A GCxGC system consists of an injector, two columns, a modulator and one or more

detectors and ovens. In 1D, conventional column dimensions (e.g. 0.25mm I.DD and 15 to

30m in length) are typically used and as mentioned in 2D shorter columns (1 – 5m) need to

be used to allow fast analysis.

The modulator in comprehensive GC can be considered the most crucial part of the

equipment, as it has three major functions: (1) continuous collection of small fractions of

effluent from 1D, ensuring separation is maintained; (2) refocussing the effluent; (3) quick

transfer of the collected 1D fraction as narrow pulse. These three processed together are a

single modulation cycle, which is repeated throughout the measurement.

Modulation can be either performed by thermal modulation or flow modulation. In thermal

modulation, the components eluting from the first dimension are immobilized by means of

liquid nitrogen. After a certain time-interval, a hot stream pulse is used to mobilize the

compounds and inject them into the second dimension. In flow modulation, differential

flows are used instead of temperature to fill and flush the sample loop of the modulator.

18

Figure 2.9: Schematic representation of a comprehensive GC system.

Due to the complementing stationary phases applied in multidimensional GC, separations

can be achieved on molecules which would generally be impossible to separate when only

one type of stationary phase would be applied. This allows for separation of more complex

samples compared to what is possible on a single column.

19

Chapter 3

Chirality and chiral separations

Since the aim of this master dissertations is to synthesize high temperature-stable stationary

phases capable of chiral separations, first the basics of chirality and chiral separations will be

explained.

3.1 Chirality

Chirality is a property of molecules lacking any symmetry elements, it refers to a molecule

which is non-superimposable on its own mirror image. The best example of a chiral object is

a pair of hands, since they both consist of the same “components”, being a thumb and four

other fingers, but they are not superimposable with their mirror image. For this reason, the

chirality of a molecule is sometimes referred to as “handiness of a molecule”.

Conventionally, mirror images of a chiral molecule (enantiomers) are designated to either

being right-handed (R) or left-handed (S). Chiral enantiomers can be distinguished by use of

polarised light, since one enantiomer will typically rotate the plane of the beam clockwise,

while the other enantiomer does the opposite.

Chirality has a major influence on the biological activity and therefor is of importance for

pharmaceutical applications since one enantiomer can have the desired pharmaceutical

effect while the other one possibly has either no effect or a detrimental one.

Due to the similarity between enantiomers, they show similar chemical and physical

properties. Therefore, separating enantiomers is more complex compared to the separation

of achiral molecules (see §3.2).

Figure 3.1: Example of two chiral molecules which can be separated by means of GC,

(R)-2-octanol (left) and (S)-2-octanol (right).

Therefor in this section the contemporary approaches allowing chiral separation of chiral

solutes by gas chromatography are discussed.

20

There are two ways in which a chiral stationary phase in GC can be obtained, either the

stationary phase can be made chiral or a certain weight percentage of chiral selector can be

dissolved into an achiral stationary phase to make it chiral.18,19

The main conventionally used stationary phases for chiral separation are based off

cyclodextrin-based phases (CD).20 These generally consist of stationary phases in which a

certain percentage of cyclodextrins are dissolved as chiral selectors. The selectivity and

sensitivity of these CD-based phases depends on the amount of dissolved selector and which

type: α-, β-, and γ-cyclodextrin, is used. Since with changing the type of cyclodextrin, the

central cavity which functions as chiral selector differs depending on the CD. (Table 3.1).21

Type of CD α β γ

Number of glucose units 6 7 8

Molecular mass (g/mol) 972 1135 1297

Inner diameter (nm) 0.57 0.78 0.95

Table 3.1: Specifications of the different types of cyclodextrins.21

As shortly mentioned above, chiral selectivity in CD-based stationary phases is based off

inclusion interactions in the central cavity of the CD, which allows for one enantiomer to be

retained better than the others.

To improve chiral separations, derivatisation of the enantiomers is also a possibility.

Derivatisation can help alter the vapour pressure, temperature stability, and interaction with

the stationary phase which all lead to improvements in terms of separation.22

Figure 3.2: Chemical Structures and sizes of three most common cyclodextrins.22

21

These phases are however usually limited by their maximum operational temperatures and

polarity. Therefore, research has been performed towards the improvement of these chiral

stationary phases, such as the development of permethylated β-cyclodextrins23,24, use of

amino acid derivatives25,26, covalent organic frameworks (COFs)27,28, and more. In more

recent years, research has been performed on the development of chiral stationary phases

based off ionic liquids (see Chapter 4) to improve thermal stabilities with the development of

e.g. an ephedrinium-based stationary phase (Figure 3.3).24 However, a lot of research is still

required before this type of stationary phases can be considered useful for commercial

applications. Hence, conventional chiral separations in GC are still limited to CD-based chiral

stationary phases.

Figure 3.3: Structure of the first used chiral ionic liquid with chiral cation

as stationary phase, the ephedrinium-ion.29

22

3.2 Thermodynamics of chiral separations

Before getting to chiral separations, first the basic principles of thermodynamics of

separations need to be explained. (Chiral) separation in chromatography is achieved by

means of partitioning of the compounds in the sample between stationary and mobile

phase. Meaning that separation solely depends on thermodynamics:25

ΔG = ΔH − TΔS (3.2.1)

Under linear chromatographic conditions, the temperature dependence of the retention of a

given analyte can be expressed by means of the van ‘t Hoff equation30:

ln 𝑘 = −

Δ𝐻°

𝑅𝑇+

Δ𝑆°

𝑅+ ln 𝜙

(3.2.2)

Where k is the retention factor, R the universal gas constant, T the absolute temperature in

Kelvin, and φ the phase ratio (𝜙 =𝑉𝑠

𝑉𝑚).31 Combination of equation 3.2.2 with the Gibbs-

Helmholtz relationship (where α is the enantioseparation factor: 𝛼 =𝑘𝑅

𝑘𝑆):

Δ(Δ𝐺) = 𝑅𝑇𝑙𝑛(𝛼) (3.2.3)

Yields equation 3.2.4:

ln 𝛼 = −

ΔΔ𝐻°

𝑅𝑇+

ΔΔ𝑆°

𝑅

(3.2.4)

The expression above relates the temperature and the experimentally accessible α-value

with the molar differential enthalpy and entropy, ΔΔH and ΔΔ, of enantioselective

adsorption. Equation 3.2.3 also shows that at lower temperatures, the effectiveness of

intramolecular interactions is increased and therefor enantioselectiveness is improved.31

As mentioned above, derivatisation can lead to better enantioselective results, which is due

to the change in interaction with the stationary phase (influencing the enthalpy factor

above) and can therefor indeed lead to better chiral separations.

23

Chapter 4

Ionic liquids

Other than the aim for making a new chiral stationary phase as mentioned at the start of

Chapter 3, it is also the goal to produce stationary phases which are both thermally stable

and have the capability to achieve separation of polar analytes. Ionic liquids are a type of

molecules which contain both characteristics. Since these are a more recent discovery, ionic

liquids and their general properties will be described here.

4.1 Introduction to ionic liquids Ionic liquids (ILs) are salts with low melting temperatures, usually below 100°C. Some ionic

liquids are even observed to be liquid at room temperature, hence also referred to as RTILs.

As their name implies, these liquids show ionic properties because they are generally built

up out of an organic cation and (in)organic anion functionality which both persist by

stabilising each other.

Such liquids show several interesting properties such as a virtually non-existent vapour

pressure, boasting a high electrochemical and chemical stability towards harsh acids and

bases, high heat capacity and as mentioned in the first paragraph, a low melting point.18

They also have the advantage of a high adaptability since they can be specifically tuned for

their intended purpose by changing the anion and cation functionalities and can be made

more thermally stable or to show different physical properties because of it. Due to these

characteristics, ionic liquids have lately become a topic of high interest in multiple branches

of chemistry such as in organic synthesis (specific chiral reactivity), organic catalysis (some

ILs offer higher yield compared to conventionally used catalysts), electrochemistry, green

chemistry (storage of radioactive materials and relative easiness to recycle), and last, but not

least: in chemical analysis.

One of such uses of ionic liquids in separation sciences is their application as stationary

phase in chromatography, whereby applications in gas chromatography are the most

promising. Ionic liquids are interesting since they show valuable properties for their use as

novel types of stationary phases. The most important properties are their polarity (due to

their ionic nature), their high thermal stability (which allows increasing the maximal allowed

24

operational temperature and thus allowing for more compounds to be measured/resolved

with GC), good capability to wet glass surfaces (giving a good coating efficiency), high

viscosity (which offers higher resistance to film disruption of the stationary phase at higher

temperatures), non-flammability, and their adequate solubility in volatile organic solvents

(which increases the ease of coating and therefore coating efficiency when coating the

columns with the static coating method).

As mentioned above, the physical and chemical properties of ionic liquids can easily be

tuned by making changes to the selected cations and anions. For example, thermal stability

of the ionic liquids can be improved by making use of a bis(trifluoromethane)sulfonimide

anion (NTf2-). Below are some of the most conventionally used anions (Figure 4.1) and

cations (Figures 4.2 and 4.3) used for ionic liquids.

Figure 4.1: Structures of conventionally used anions for ionic liquids bis(trifluoromethane)sulfonimide, NTf2-

(left), hexafluorophosphate (middle) and halides (right).

Figure 4.2: Structures of “onium” based cations for ionic liquids, phosphonium (left) and ammonium (right).

Figure 4.3: Typical cations pyridinium (left) and imidazolium (right).

25

Imidazolium-based ionic liquids are conventionally named as [R1R2Im] where R1 and R2 are

the two organic groups bound to the nitrogen functionalities of the imidazolium-ring as

represented in Figure 4.3.

Since the organic chains on ionic liquids mainly consist of simple alkanes, naming of the

different cations is done by shortening the chain present on the “onium” moiety to either

the first letter (e.g. M for methyl, B for butyl, …) or writing down their chain length (e.g. C12

for dodecane, …).

4.2 Development of IL stationary phases

The first research performed on using molten salts in GC started in 1959, when Barber et al.

synthesized stationary phases based on stearate salts of bivalent metals (manganese, cobalt,

nickel, copper, and zinc)29,32. During the 1980s, Poole et al. synthesized ethyl ammonium

nitrate and ethyl pyridinium bromide stationary phases, which were capable of being used as

polar stationary phases33. Along with these stationary phases, alkyl ammonium- and alkyl

phosphonium salts were also synthesized. These early IL-based stationary phases, however,

suffered from narrow liquid ranges, low column efficiencies and/or from poor thermal

stabilities, which meant that they were difficult to use as stationary phases.

Figure 4.4: ethylammonium nitrate (left) and ethylpyridinium bromide (right) salts used in

the early research in the field of IL-based stationary phases.

It was not until 1999 that the group of Armstrong managed to synthesize ionic liquids which

were usable as stationary phase3 ([BMIM][Cl] and [BMIM][PF6]). It’s with these imidazolium-

based stationary phases that the “dual nature” of ILs as stationary phase was observed, since

they had the capability of separating polar and apolar compounds as if they were

respectively polar and apolar stationary phases.

26

Figure 4.5: Structures of [BMIM][Cl] (left) and [BMIM][PF6] (right).

In 2001, the group of Armstrong went even one step further and attempted at producing a

chiral IL-based stationary phase by dissolving methylated cyclodextrins in the [BMIM][Cl]

stationary phases they synthesized back in 1999.34 The IL appeared to be a useful stationary

phase for GC and it was able to dissolve more than 25% w/w without intervening too much

with the retention on the column. However, the cyclodextrins used formed inclusion

complexes with small aromatic molecules and with the [BMIM][Cl], which hindered the

chiral separation of these stationary phases based on the inclusion mechanism. It was also

found that [BMIM][Cl] begins to degrade at a temperature of 120°C by starting to show

discolouration, with obvious signs of decomposition starting at 150°C. Which limits the

maximum temperature at which this stationary phase can be used for GC to 120°C.35

Only two years later, in 2003, Armstrong and co-workers managed to synthesize new IL

stationary phases which had increased thermal stability compared to [BMIM][Cl]. With

thermal stability up to around 250°C for 1-(4-methoxyphenyl)-3-methylimidazolium tri-

fluoromethanesulfonate ([MPMIM][TfO]). They also observed that the anion has some effect

on the separation efficiency when separating molecules capable of donating protons.36

In 2004, also the group of Armstrong managed to synthesize the first chiral IL capable of

performing chiral separations. The N,N-dimethylephedrinium-based chiral stationary phases

they synthesized allowed for effective separation of enantiomers of alcohols (Figure 4.6),

diols, sulfoxides and some N-blocked amines and epoxides. However, it was observed that

after several weeks of use at temperatures above 140°C, their stationary phase lost

enantioselectivity for certain compounds but not for others. Which they contributed to

dehydrogenation of the hydroxyl group (one of the two stereogenic centres). Since

hydrogenation, followed by the addition of water would induce racemisation at this centre.37

27

Figure 4.6: GC chromatogram of chiral separation of (left to right), sec-phenethyl alcohol, 1-phenyl-1-butanol,

and trans-1,2-cyclohexanediol. On a column (8m long, 250µm internal diameter) coated with (1S,2R)-(+)-N,N-

dimethylephedrinium-bis(trifluoromethanesulfon)imidate.29

In 2005, Anderson and Armstrong attempted to synthesize a number of cross-linked achiral

ionic liquid stationary phases.38 These di-cationic cross-linked ILs based on imidazolium and

pyrrolidinium cations were characterised later that year by the group of Anderson. The

following trends were observed in the di-cationic ILs synthesized:39

• Longer linkage chains between cations decrease the melting point

• Density of ILs decreases with increasing linkage chain length

• Length of the substituted alkyl-group on the imidazolium ring decreases surface

tension with increasing length

• A longer alkyl group at the three position of the imidazolium ring increases viscosity

In the subsequent years, efforts were made at increasing the column operation temperature

and increase the selectivity ranges by developing di-cationic ILs, functionalized ILs,

polymerized ILs and even producing stationary phases by mixing ILs. This caused a major

increase in the range of possibilities for ILs as stationary phases since they were previously

limited by either thermal stability or selectivity.

28

Due to the increased interest of usage of ILs in separation sciences, research was performed

by the group of Lynen in 2016 to investigate the influence of polymerisation mechanism on

the thermal stability of polymeric ionic liquids (PILs).4 By comparing chain-growth PIL

stationary phases to step-growth (condensation polymerisation) ones, they discovered that

chain-growth PILs perform significantly better than their step-growth (radical

polymerisation) counterparts in terms of thermal stability, column efficiency, minimal plate

heights, and retention and peak shape of polar solutes. There was also no significant

drawback of these PIL stationary phases when compared to commercial non-polymerised

ionic liquid-based columns, while increasing the thermal stability.

Recently, the attention of ILs in separation sciences seems to have shifted from development

of new IL stationary phases towards the use of ionic liquids for multi-dimensional GC

measurements40 and analysis of more complex samples such as fatty acid methyl esters

(FAMEs).41 Some of these ionic liquids have also been more used for e.g. extractions such as

in solid-phase micro-extraction (SPME).42–46

Some of the stationary phases reported above have also become commercially available.

These commercially available columns have stationary phases which are based on the di-

cationic linked stationary phases synthesized by Armstrong and Anderson in 2015.

These stationary phases are mainly being sold by Sigma Aldrich under the name of patented

Supelco® SLB-IL series columns of which they claim these can be engineered with either

identical selectivity to non-ionic liquid columns but with higher resistance to damage due to

moisture/oxygen and higher operation temperatures or completely unique selectivity

compared to their non-ionic liquid counterparts. These columns are also suggested for their

use in multidimensional separations. The chemical structures of some of these Supelco® SLB-

IL columns can be found below in Figure 5.5. The naming of the columns is based on polarity,

with the number after the SLB-IL pointing out the polarity of the column.

29

Figure 4.7: Two commercially available IL-based stationary phases phosphonium-based SLB-IL59 (left)

and imidazolium-based SLB-IL111 (right)41,47

4.3 Applications of IL stationary phases

Since the section above deals with developments of new stationary phases and only shortly

lists their advantages and main findings, it doesn’t give much insight into the advantages and

drawbacks of IL stationary phases compared to the conventionally used stationary phases

such as wax and PDMS columns. Hence the importance of testing the applicability of the

stationary phases.

A lot of the commercially available IL-based stationary phases, such as those synthesized by

the group of Armstrong et al. were tested on a variety of different samples. However, it is

the group of Mondello et al. who investigated the applicability of these stationary phases by

measurement of complex samples in comprehensive GC (GCxGC). These tests are significant

since they show the strenghts and drawbacks of these kinds of stationary phases when

applied to less ideal samples which are generally obtained when performing routine

measurements.

An interesting set of practical tests of such commercially available IL-based stationary phases

was performed by Ragonese in 2009 who managed to separate fatty acid methyl esters

(FAMEs) in diesel blends by means of a 1,9-di(3-vinyl- imidazolium)nonane

bis(trifluoromethyl)sulfonylimidate stationary phase (SLB-IL100).48 The main progress

offered by these columns is the fact that separation of hydrocarbons from the target

compounds was possible within a complex sample such as diesel, which was possible

without requiring complex sample preparation or the use of multidimensional techniques,

which are generally required for such samples.

30

The observation was made that with respect to stationary phases generally used for these

kinds of samples, an IL-based column with the same dimensions was capable of performing

the same analysis in a reduced time. By plotting the carbon numbers of the FAMEs against

elution temperatures, the results indicated that the highest Cn-values obtained with the

conventionally used polyethylene glycol stationary phases would be C28, whereas the longest

chain that could potentially elute fro mthe IL column at its MaOT would be C32. Which would

increase the amount of components in FAME mixtures that can be measured my means of

gas chromatography, broadening the field for measurements in the field of e.g. biodiesel

analysis.

In 2011, a comparison was made between the power of ionic liquid columns with the

separation power of stationary phases containing 100% polyethyleneglycol. It was

demonstrated that the ionic liquid columns were a valuable tool for achieving excellent

separation performance in the field of flavor and fragrance as well. The conclusion was even

made that in term of the overall observed performance for the analysis of essential oils, the

IL phase performed better than the conventionally used wax (100% polyethylene glycol)

columns in both identification and quantification when applied in GC-MS measurements.49

The power of the IL stationary phases was also shown in 2011, when these columns were

applied for the first time in hyphenated gas chromatography techniques such as GCxGC50 by

the separation science group . The main advantage of these ionic liquid stationary phases are

their flexible selecitivities and high thermal stabilities which might make these stationary

phases interesting not as replacement of the conventionally used columns in these

techniques but more as an addition to the already existing ones.51 This was later followed by

similar shorter broadening the scope of GCxGC application.

The applications above will likely increase as more time and development is put into the

research of the performance of new ionic liquid stationary phases.

31

The goal of this master dissertation is to further improve upon the applications mentioned

above by synthesis of new polymeric ionic liquid stationary phases capable of chiral

separation. The attempt will be made to achieve chiral separation by synthesizing an ionic

liquid which is chiral itself without having to rely on addition of chiral selectors. These

stationary phases will be synthesized by starting from basic molecules, followed by a coating

step to apply them to a column. These columns will then be tested for thermal stability and

retention and selectivity investigation to be able to properly determine the characteristics of

these new stationary phases.

32

33

Chapter 5

Synthesis and application of chain-grown polymeric

ionic liquids

To obtain a chiral stationary phase, the first attempt was performed by synthesis of a chain-

growth polymeric ionic liquid (PILs). This was mainly because the synthesis of similar PILs

were already performed and reported by Roeleveld et al.4 It also served as a good starting

point for this thesis since the synthesis and purification required were rather simple.

5.1 Synthesis of chain-growth monomer

5.1.1 Monomer synthesis

The monomer required for the chain-growth polymerisation was synthesized by adding 8mL

of (S)-(+)-1-bromo-2-methylbutane to a solution containing 5mL 1-vinylimidazole in 20mL of

dry ethyl acetate. After adding both products together, the reaction was then heated to 77°C

to reflux and left for 48h to obtain as much product as possible.

Figure 5.1: Synthesis route for the chain-growth monomer

At the end of the reaction, a separate viscous layer formed at the bottom of the flask since

the monomer was no longer soluble in ethyl acetate, which allowed for an easy purification

of the desired product by means of decanting and washing the product with dry ethyl

acetate to remove unreacted reagents still present in the flask. Since after the reaction,

some reagents proved still present, approximately 5mL of dry ethyl acetate was added to the

solution and the mixture was put into an ultrasonic bath for 30min to allow thorough access

for the solvent into the viscous product. In this way of purification was repeated until the

fraction of 1-vinylimidazole and (S)-(+)-bromo-2-methylbutane still present in the product

was negligible.

34

5.1.2 Polymer synthesis

For the polymer synthesis, the same procedure was followed as by Roeleveld et al..4 In which

the chain-growth was performed by means of a radical polymerisation.

4.040g of the monomer obtained in §6.1 was dissolved into 15mL dry DMF and to this

monomer 2mol% of AIBN (0.33mmol) was added as initiator. The solution was then purged

with argon to remove traces of oxygen to avoid unwanted transfer- and termination

reactions, as oxygen is a good transfer-reagent in radical polymerisation. After purging, the

reaction mixture was then heated to 70°C and left to polymerise overnight (~12h).

Figure 5.2: Synthesis route for the polymerisation of the chain-growth PIL, CPIL1

No separate polymer layer was observed after the reaction, so the flask was put in an ice

bath and the polymer was precipitated by adding acetone, after which the polymer

precipitated as a white powder. This white powder was then simply purified by filtering off

over a sintered glass filter and washing with acetone, after which the polymer remained on

the filter, was collected into a vial and put in a vacuum oven to dry. The obtained polymer at

this point was named CPIL1.

35

5.1.3 Ion exchange reaction

Due to the limited thermal stabilities of imidazolium-based ILs with halide anions reported in

literature 52, the anion of CPIL1 was exchanged with a more temperature stable NTf2—anion.

The reaction was performed as follows, a 2g of CPIL1 obtained in §5.1.2 was dissolved into

7mL water and subsequently a solution containing 3 equivalents of LiNTf2 (6.959g) in 7mL

water was added to ensure the presence of plenty of exchange reagent. The mixture was

then left to mix for a day after which a separate white layer could be observed.

Figure 5.3: Reaction scheme for the ion exchange of CPIL1

The precipitate was then filtered off over a porous glass filter and washed with water until

no more bromine was detected in the water by means of AgNO3. The resulting product was

named CPIL2 and was put into a vacuum oven to dry overnight.

5.2 Coating of chain-growth ionic liquid polymer The obtained ionic liquid polymers were coated on 5m and 30m columns by means of the

static coating method (Figure 5.4). When coating a column through static coating, polymer is

brought into solution, after which it is sent through the column. After the column is filled,

the one end of the column is sealed off, while the other is connected to a vacuum pump, the

entire column is then submerged into a temperature heated water bath to evaporate the

solvent in which the polymer was dissolved to remove it and deposit a homogeneous film of

polymer on the inside of the column.

The thickness of the deposited film can be varied by making use of Equation 5.2.1:

𝑐 =

4𝑑𝑓𝜌

𝑑𝑐 (5.2.1)

36

In which df is the thickness of the film, ρ is the density of the polymer (in g/cm3), and dc is the

inner diameter of the column. Since time limitation didn’t allow to perform density

measurements on the obtained polymers, the densities of CPIL1 and CPIL2 were assumed to

be the same due to them only having differing anions. The density was estimated to be close

to 0.7019g/cm3, due to the high similarity to the ionic liquids synthesized in the work of

Roeleveld et al..4

Hence, for the coating a solution was made of 4mg/ml in DCM and acetone for CPIL1 and

CPIL2 respectively. Desired lengths for the column were taken from an untreated silica

capillary GC column with an internal diameter of 0.25mm, which with the concentration

above would lead to a thickness of film of 0.35µm.

Since the starting columns used for coating were untreated, surface treatment and

optimisation steps had to be performed first. For this, an NaOH solution with a

concentration of 1M was flushed through the column with the use of N2 pressure (see Figure

5.4) after which the column was rinsed by means of Milli-Q water. Next up, a HCl solution

with a concentration of 1M was sent through the column after which it was rinsed again

with Milli-Q water. Then, MeOH was flushed through to remove traces of water.

The coating of the polymer was then performed by first flushing the solvent in which the

desired stationary phase IL was dissolved (DCM for CPIL1 and acetone for CPIL2) after which

the solution containing the polymer was sent through the column. When the capillary was

filled with polymer solution, the nitrogen pressure was lowered to atmospheric pressure and

the end of the column was submerged into silicone while the polymer solution still dripped

from the column. This sealing step is the most crucial one in the coating process as the

presence of the slightest amount of air at the end of the column would lead to the column

being emptied of the polymer solution before the polymer can be deposited on the inner

walls of the column when applying the vacuum.

After leaving the end of the column in the silicone for 30min to obtain proper sealing, the

other end was attached to a vacuum pump and the entire column was submerged into a

temperature-controlled water bath which was set at 25°C for CPIL1 and 40°C for CPIL2 to

evaporate the solvent.

37

Figure 5.4: Representation of the static coating method steps: pre-treatment and column filling step (left),

solvent evaporation step with deposition of stationary phase (right)

After coating, the columns were conditioned for two hours at 120°C in a GC-FID under a

hydrogen flow of 1.2ml/min before performing any measurements to increase the longevity

of the stationary phase. The columns were then tested by means of two test solutions. One

containing two alkanes (dodecane and pentadecane) in a concentration of 100ppm, the

other containing benzaldehyde, 3-nitrotoluene, and benzyl benzoate, also in a concentration

of 100ppm. The reason why these samples were chosen is due to the fact that for similar

imidazolium-based ionic liquid stationary phases, poor retention for alkanes was reported

whereas highly polar compounds such as benzaldehyde offered good peak shapes and

retention.4

The solution of alkanes was injected to check if coating was present, since presence of the

coating would lead to poor peak shapes and retention due to interactions with the

stationary phase. Lack of coating would lead to good peak shapes due to separation only

being achieved based on differences in boiling point.

Upon passing this initial test, further measurements were performed on the columns

whereas upon failure, another column had to be coated by means of the static coating

approach.

38

5.3 Results and discussion

5.3.1 Assessment of thermal stability

The thermal stabilities of CPIL1 and CPIL2 were assessed by means of TGA analysis. For this

respectively 2.44 mg and 4.82 mg of CPIL1 and CPIL2 were taken and placed on a platinum

plate connected to a balance. Subsequently, the oven was heated from 25°C to 800°C at

10°C/min under air while the weight loss was followed as function of the temperature

(Figure 5.5).

Figure 5.5: TGA measurement of CPIL1 and CPIL2

As can be observed in the graph above, an initial weight loss was detected for CPIL1 below

100°C. This is likely due to the evaporation of remains of water present in the polymer. The

first significant weight loss detected for CPIL1 was at 250°C which was determined to be the

onset degradation temperature, Td.

The first weight loss for CPIL2 was detected at a temperature of approximately 300°C, which

was determined to be the onset degradation temperature, Td, for CPIL2.

It is clear from the TGA measurement that CPIL2 has a higher onset degradation

temperature compared to CPIL1. Which is in line with earlier findings reported in literature,

which already found that the thermal stability of an IL increased by approximately 50°C after

exchanging a halide anion by a more thermally stable bis-(trifluoromethane) sulfonimide,

NTf2-, anion.

39

Another interesting aspect which is clarified by the TGA measurements of CPIL1 and CPIL2 is

that CPIL1 is likely hygroscopic by nature due to the detection of an initial weight loss around

100°C, whereas this weight loss was not detected on the measurement of CPIL2. This would

mean that CPIL2 is likely not hygroscopic.

TGA measurements however are incomplete to describe the properties of CPIL1 and CPIL2

when applied as stationary phases in GC, since it is only sensitive to volatilisation of the

sample in question whereas at temperatures below the onset degradation temperature,

degradation of the stationary phase might already be possible, which would cause e.g.

disruptions in the homogeneity of the stationary phase.

Therefore, CPIL1 and CPIL2 were also tested in bleed tests after they were coated using the

static coating method (see §5.2). For these tests, columns with the following dimensions

were used 5m x 0.25mm x 0.35µm. The measurements were conducted under a H2-flow of

1mL/min and using a temperature program starting at 40°C to the maximum applied

temperature (200°C – 275°C) at a rate of 10°C/min, after which the maximum temperature

was held for another 10min. In these bleed measurements, the increase of the background

was followed, and the maximum allowed increase of the background was determined to be

20pA, since any column bleed above this temperature is to be avoided to increase the

lifetime of the columns.

40

Figure 5.6: Bleeding profiles of 5m columns coated with CPIL1 and CPIL2 for a blank analysis on GC-FID with a

temperature program starting at 40°C and ending at different maximum temperatures held for 10min.

From the chromatograms of the bleed measurements, which can be observed above in

Figure 5.6, the maximum operational temperature (MaOT) of columns coated with either

CPIL1 or CPIL2 can be determined. This MaOT value is the highest temperature for which the

“bleed” or background of the column does not increase more than 20pA. Since a background

signal higher than this value would imply detrimental degradation of the stationary phase in

the column.

The bleed measurements have shown that the MaOT of CPIL1 is around 200°C, since at

higher temperatures, the background already goes up to 160pA. Whereas the MaOT for

columns coated with CPIL2 goes up to close to 250°C.

When comparing the results of the bleed measurements with the initial assessment on

thermal stability with use of TGA, the same trend is obtained in which the thermal stability

of CPIL2 is approximately 50°C higher than the one for CPIL1 although, it should be noted

that the actual maximum temperature at which the columns can be used is approximately

41

50°C lower than the values obtained from the TGA measurement. This is most likely due to

the increased sensitivity of an FID-detector in comparison to a mass balance. Nevertheless,

the conclusion can be made that the use of TGA measurements to assess thermal stabilities

offers an overestimation on the maximum temperature which can be applied for a column

used in GC.

5.3.2 Evaluation of retention and separation

As mentioned before in §6.2, an initial test was performed on the polymers. After passing

this test, further assessment on retention and separation was performed. Due to the limited

thermal stability of CPIL1, no further measurements were performed on this column as it

was considered inferior to CPIL2 in terms of thermal stability.

The column coated with CPIL2 was tested for its retention and separation characteristics by

measuring a test mix containing five polar compounds: methylsalycylate, benzaldehyde,

benzyl benzoate, ɣ-nonanoic lactone, and 3-nitrotoluene. All in a concentration of 100ppm,

using the experimental parameters found in Table 5.2:

Inlet Temperature 200°C

Detector Temperature 200°C

Flow (H2) 1.2ml/min

Average Velocity 41cm/s

Split ratio 2:1

Temperature program 40°C – 225°C at 2.5°C/min

Table 5.2: Experimental parameters for retention and separation tests of CPIL2

The following chromatogram was obtained (Figure 5.7), which showed proper separation of

the five test compounds although low signal intensities were observed due to a significant

broadening of the peaks.

More bleed was observed in the chromatograms below, which caused by the fact that a

longer column was used for the retention measurements.

42

Figure 5.7: Chromatogram for sample containing 100ppm of benzaldehyde (A), methylsalycylate (B),

3-nitrotoluene (C), ɣ-nonanoic lactone (D), and benzyl benzoate (E) measured on a column coated with CPIL2

(30m x 0.25mm x 0.35µm)

Proper separation was achieved, although it was noted that the peaks observed for

3-nitrotoluene, benzyl benzoate and ɣ-nonanoic lactone were significantly sharper than the

peaks obtained for benzaldehyde and methylsalycylate.

To broaden the evaluation of the retention and separation characteristics of the CPIL2

stationary phase, a sample was made containing 100ppm of acetophenone, propiophenone,

benzophenone, butyrophenone, and hexanophenone. In this way it could be assessed how

closely related compounds would be retained on the stationary phase.

Figure 5.8: Chromatogram for sample containing 100ppm of acetophenone (A), propiophenone (B),

butyrophenone (C), benzophenone (D), hexanophenone (E) measured on a column coated with CPIL2

(30m x 0.25mm x 0.35µm)

A B C

D

E

C A B

E

D

43

Separation was achieved for all compounds present, however no baseline separation was

achieved for acetophenone and propiophenone, likely due to the high similarities between

both molecules.

It was noted that for the two samples measured above (Figures 5.7 and 5.8) the same order

of retention was observed as for the stationary phases synthesized by Roeleveld et al.4

5.3.3 Assessment of column efficiency

The general way to assess column efficiency is to determine the minimal plate height

through Van Deemter curves. Due to the limited thermal properties of CPIL1, however, these

measurements were only performed on CPIL2.

For this, two probe compounds from the evaluation of retention, benzyl benzoate and ɣ-

nonanoic lactone were prepared in a concentration of 100ppm. These components were

measured at a temperature of 110°C and at a split ratio of 10:1, whereas the average

velocity over the column was varied from 20cm/s to 100cm/s, as 20cm/s was the lowest

velocity that could be applied for the GC-FID and the amount of plates in the chromatogram

were determined at each velocity. Measurements were performed in threefold to obtain a

reliable result for the plate counts.

Dividing the values in the table above by the length of the column (5m), the plate height can

be set out as a function of the average velocity of the mobile phase. Plotting these values,

yields the Van Deemter curves for these two compounds on CPIL2 (Figure 5.9)

44

Figure 5.9: Van Deemter curves for ɣ-nonanoic lactone (black) and benzyl benzoate (red) on CPIL2

measured isothermally at 110°C

The minimal obtained plate height for ɣ-nonanoic lactone was determined to be 0.99mm at

the optimal mobile phase velocity of 30cm/s, whereas for benzyl benzoate the minimal plate

height was 1.42mm with an optimal mobile phase velocity of 25cm/s.

To fully assess the column efficiency, the coating efficiency can be determined with use of

the formula below:

𝐶𝑜𝑎𝑡𝑖𝑛𝑔 𝑒𝑓𝑓𝑖𝑐𝑖𝑒𝑛𝑐𝑦 (%) =

𝐻𝑚𝑖𝑛(𝑇ℎ𝑒𝑜𝑟𝑒𝑡𝑖𝑐𝑎𝑙)

𝐻𝑚𝑖𝑛(𝐸𝑥𝑝𝑒𝑟𝑖𝑚𝑒𝑛𝑡𝑎𝑙) (5.3.1)

The theoretical minimal plate height can be determined by making use of Equation 2.3.14

(Page 10), meaning that the theoretical minimal plate height would be equal to 0.25mm.

Which leads to a value for the coating efficiency of only 25.25% when using the lowest plate

height obtained in the measurements above. Meaning that the coating on the column is only

a quarter as efficient as it should be considering theoretical values. Although, the deviation

from these values are likely since the thickness of the film applied to the column is 0.35µm,

which cannot be considered as a thin coating anymore in terms of the derivation used to get

to Equation 2.3.14. Even so, the coating efficiency is still rather low, which means there is

still room for improvement in terms of coating and purity of the polymer, since increased

purity will cause a better film homogeneity, which leads to increased coating efficiency.

45

5.3.4 Assessment of chiral separations

Measurements to check the capability of chiral separations were performed on CPIL2.

However, no chiral capabilities of this stationary phase were observed. There are three

possible reasons which could explain this issue.

The first reason would be the fact that due to polymerising the chiral monomer by means of

radical polymerisation possibly could have racemised the polymer.

Secondly, it could simply be that having a methyl group as only chiral centre in the polymer

could simply not suffice to achieve chiral separations, although some chiral separation would

be expected if this was the only case, since the ephidrinium-based IL synthesized by the

group of Armstrong still showed (limited) chiral properties after the racemisation of the

chiral hydroxyl-group when applying a temperature above 140°C;37 leaving the chiral

separations to be achieved by only the methyl group. Which means that chiral separations

with only a methyl group as chiral selector should still be possible.

Finally, another viable reason that chirality was not obtained with the CPIL2 stationary phase

could be since after the polymerisation, the methyl groups are only present on the side-

chains of the polymer, which could cause a shielding effect for analytes when interacting

with the stationary phase, which would indeed lead to the lack of any capabilities of chiral

separations.

Figure 5.10: Chromatogram of attempt at chiral separation on CPIL2 (5m x 0.25mm x 0.35µm) of a racemic

mixture of carvone (left; A) and trans-1,2-cyclohexanediol (right; B)

A B

46

None of these reasons above could be tested however, although the shielding effect and

racemisation due to the way the polymer was obtained (radical polymerisation) could be

investigated upon by testing out a column coated with CPIL3 which is discussed further.

5.3.5 Influence of thickness of coating

The influence of the thickness of coating was determined by coating a 5m column by using a

solution at a concentration of 3mg/ml, which yielded a column with a thickness of film of

0.25µm. On this column, the same sample was measured as mentioned above (§5.3.2) and

the efficiency of this column was compared to the measurement on the column with a

coating of 0.35µm by a comparison of peak capacities (§2.5; Page 16) of the columns, since

both measurements were run under the same instrumental parameters. Since the

measurement ran from 40°C to 200°C under a gradient of 2.5°C/min, the gradient time was

determined to be 64min. Which allowed the peak capacities to be calculated as:

0.25µm coating 0.35µm coating

Peak capacity 102.175 94.227

Table 5.3: Peak capacity for CPIL2 coating of 0.25µm and 0.35µm

The peak capacity measurement above makes clear that the thinner coating of 0.25µm

would obtain better results when applied in gradient analysis when compared to a film

thickness of 0.35µm for CPIL2. In the chromatogram measured with the column containing

the 0.25µm thick coating, a significant increase of the bleed and loss of retention for the

compounds were observed.

Figure 5.11: Chromatograms showing separation of benzaldehyde (A), 3-nitrotoluene (B),

and benzyl benzoate (C) on a column coated with CPIL2 with film thickness of 0.25µm (left) and 0.35µm (right)

A

A

B

B

C

C

47

5.3.6 Comprehensive GC measurements (GCxGC-FID)

Further tests were also performed to see if the columns obtained in this work could be

applicable as second dimension in comprehensive GC measurements. For this,

measurements were performed with a second dimension CPIL2 column with a thickness of

film of 0.35µm. Initially a Zebron 5HT-inferno column (15m x 0.25mm x 0.10µm) was used

for the measurements.

Initial measurements with a 5m column as second dimension clearly showed issues with

wrap-around, meaning that analytes remain in the column for too long which causes the

signal to be completely spread over the second dimension, yielding a chromatogram

showing vertical bands since separation was still achieved over the first dimension.

To reduce the spread of the analyte over the second phase due to wrap-around, the column

in the second dimension was shortened from 5m to 2m since this way, (excessive) retention

is reduced. After shortening of the second phase, a better result, in which the analyte didn’t

spread completely over the second phase, was obtained.

Since further attempts at optimisation didn’t yield increase of results, the first dimension

was exchanged for a HP5-MS column (30m x 0.25mm x 0.25µm) since stationary phase

mismatch between the Zebron 5HT column and the CPIL2 coated column was suspected.

With the same instrumental parameters, the results improved significantly, which was

enough proof that the CPIL2 stationary phase also has some applicability in multidimensional

GC measurements as second dimension. (Figures 5.12 and 5.13)

48

Figure 5.12: Contour plot of 2D GCxGC-measurement of a mixture containing 5 compounds: acetophenone (A),

propiophenone (B), butytophenone (C), hexanophenone (D), and benzyl benzoate (E) separated on a HP5-MS

(30m x 0.25mm x 0.25µm) column as first dimension and CPIL2 (2m x 0.25mm x 0.35µm) as second dimension.

Figure 5.13: Contour plot of 2D GCxGC-measurement of a mixture containing 8 compounds: toluene (A),

anisole (B), aniline (C), butylphenylether (D), methylsalycylate (E), 3-nitrotoluene (F), cinnamaldehyde (G),

and ɣ-nonanoic lactone (H) separated on a HP5-MS (30m x 0.25mm x 0.25µm) column as first dimensional

column and CPIL2 (2m x 0.25mm x 0.35µm) as second dimension.

B

A

C D E

A B

C

D

E

F

G

H

49

Chapter 6

Synthesis and application of step-growth polymeric

ionic liquids

Since the chain-growth polymer didn’t allow to achieve results in terms of chiral separations,

another type of stationary phase was also synthesized, whereby a step-growth

polymerization process was intended. This phase could be interesting since in this way, it

could be investigated if CPIL2 didn’t manage to achieve chiral separation due to either

racemization or the chiral selector not being present on the backbone of the polymer.

6.1 Synthesis of step-growth polymer

6.1.1 Monomer synthesis

The monomer required for the step-growth polymerisation was synthesized by slowly adding

a solution containing 0.975g imidazole in 5mL dry THF to a suspension of 0.129g LiH in 10mL

dry THF while stirring and cooling in an ice bath to 0°C. After 90min, 1.55mL of R-(-)-bromo-

2-methyl-1-propanol was added dropwise while the solution was cooled to 0°C. Next, the

mixture was left to stir for 24 hours at room temperature.

Figure 6.1: Synthesis route for the step-growth monomer

After 24h, the reaction was quenched by adding 30mL Milli-Q water and the mixture was

then extracted four times with 30mL of DCM, after which the organic fraction was dried by

adding Na2SO4. This fraction was then filtered off and concentrated at a rotary evaporator

until only a liquid oily substance remained.

50

After this purification step, imidazole was still present in the sample, so further purification

had to be performed. For this, a silica column was set up in which a solution of acetonitrile

and dichloromethane in a ratio of 2:1 was used to elute the product from the column. Since

the product remained on the column after taking over hundred fractions, the column was

flushed by adding methanol. The methanol fraction was then concentrated at a rotary

evaporator and an NMR sample was taken for full analysis to determine both the purity and

if the desired product was formed (see Appendix; Page IV).

Since for the step-growth polymerisation, a leaving group on the end of the alkane is

required, the alcohol functionality needs to be exchanged for a chlorine. This was performed

by dissolving 0.390mg of the product of the first reaction (Figure 6.1) in 5mL dry DCM after

which 2 equivalents of dry triethylamine (465µl) were slowly added. The mixture was then

cooled down to 0°C before slowly adding 2 equivalents of thionyl chloride (300µl). The

reaction was then left to stir for 24h.

For the purification step, the product was extracted threefold with 15mL of an aqueous

solution containing 10% HCl to attempt to remove the triethylamine present from the DCM

phase by protonating it. However, the product remained in the water phase. Therefore, the

water fraction was neutralised by means of a NaOH solution and subsequently concentrated

at a rotary evaporator. Due to the neutralisation above, a lot of salt was present in the vial,

therefor acetone was added. The resulting mixture was then filtered off over a porous glass

filter and washed with acetone. The acetone fraction was concentrated at a rotary

evaporator. An NMR sample was taken from the acetone fraction in D2O, which confirmed

the product was present in this fraction without any major impurities.

51

6.1.2 Polymer synthesis and ion exchange reaction

Since the polymerisation is step-growth process, there are no impurities caused by the

reaction. Therefore, the product obtained from the monomer synthesis (~50mg) was

dissolved into 1.5mL of dry DMF in a microwave vial, flushed with argon to remove possible

traces of water. After flushing with argon, the reaction was heated to 110°C and left to react

for 24h.

Figure 6.2: Synthesis route for the polymerisation of the step-growth monomer

After the polymerisation, acetone was added in excess to induce precipitation. Because it

was noted that the precipitate stuck to the glass completely, to increase the yield, the vial

was cooled in an ice-bath after which the solution was centrifuged.

The acetone was then decanted off and the vial was concentrated with use of a rotary

evaporator to remove remaining traces of acetone. Chloroform was then added to remove

remaining traces of monomer. The chloroform was decanted off and the solution was dried

at a rotary evaporator.

To start the ion exchange reaction, the resulting product (~40mg) was dissolved into 2.5mL

of water and the polymer solution was transferred back to a microwave vial. An excess of

LiNTf2 (727mg) was added to ensure enough reagent for the exchange reaction and the

reaction was then stirred for 24h.

Figure 6.3: Synthesis route for the ion exchange procedure

Because the polymer containing the NTf2-anion was no longer soluble in water, purification

was performed in a straight-forward manner by simply decanting and washing until no more

chlorine was detected in the decanted water fraction with means of AgNO3. The resulting

product was then put into a vacuum oven to dry.

52

6.2 Coating of step-growth polymer

The coating of the step-growth polymer was also done by means of the static coating

method (§5.2; Pages 35-37). As was the case for the chain-growth based stationary phases,

the density was assumed to be 0.7019g/cm3. Therefor a solution of CPIL3 in acetone was

made at a concentration of 4mg/ml to obtain an estimated coating thickness of 0.35µm.

6.3 Results and discussion

6.3.1 Assessment of thermal stability

Due to time-limitations, the thermal stability of CPIL3 was only assessed by means of TGA

analysis. For these measurements, 4.053mg of CPIL3 was put onto a balance and the weight

of the polymer was measured under air while temperature was increased from 25°C to

800°C at a gradient of 10°C/min.

Figure 6.4: TGA measurement of CPIL3

As can be observed from the graph of the TGA data (Figure 6.4), there is an initial loss of

weight noticeable at around 160°C, which could be due to a fraction of monomer still being

present in the sample. From the TGA measurement, the onset degradation temperature of

CPIL3 was determined to be 300°C.

Which would mean that the maximum operational temperature of CPIL3 as stationary phase

is likely 250°C, which would make it as stable as CPIL2. Although, further research is required

on the thermal stability of CPIL3.

53

6.3.2 Evaluation of retention and separation

The retention and separation characteristics of CPIL3 were assessed by means of two test

solutions. One containing three polar compounds in a concentration of 100ppm:

benzaldehyde, 3-nitrotoluene, and benzyl benzoate. The other being the phenone mix which

was also used to assess separation and retention of CPIL2, containing acetophenone,

propiophenone, butyrophenone, benzophenone, and hexanophenone in concentrations of

100ppm each.

The measurements were performed using the experimental parameters found in Table 6.1

Inlet Temperature 200°C

Detector Temperature 200°C

Flow (H2) 1.2ml/min

Average Velocity 41cm/s

Split ratio 2:1

Temperature program 40°C – 200°C at 5°C/min

Table 6.1: Experimental parameters for retention and separation tests of CPIL3

Figure 6.5: Chromatogram for sample containing 100ppm of benzaldehyde (A),

3-nitrotoluene (B), and benzyl benzoate (C) measured on a column coated with CPIL2 (30m x 0.25mm x 0.35µm)

A

B

C

54

Figure 6.6: Chromatogram for sample containing 100ppm of acetophenone (A), propiophenone (B),

butyrophenone (C), benzophenone (D), hexanophenone (E) measured on a column coated with CPIL3

(5m x 0.25mm x 0.35µm)

The same retention characteristics are observed as for the CPIL2 stationary phase, in which

the compounds consisting of more aromatic groups were more thoroughly retained on the

column and yielded sharper peaks than the other measured compounds.

It also must be noted that for the sample containing various phenones, acetophenone and

propiophenone co-eluted completely and no baseline separation was achieved between this

co-eluting peak and the peak of butyrophenone.

Comparing the chromatograms above with the chromatograms obtained with CPIL2 as

stationary phase, leads to the conclusion that in terms of separation, CPIL3 seems to

perform worse compared to CPIL2.

since for the sample containing various phenones, no separation at all is observed between

the first three components (acetophenone, propiophenone, butyrophenone), whereas CPIL2

did manage to achieve some separation albeit no baseline separation.

A+B C

D

E

55

6.3.3 Assessment of chiral separations

The capability to achieve chiral separations with CPIL3 was tested by injecting a racemic

mixture containing two different chiral molecules which also showed good retention on

CPIL2: carvone and 1,2-transhexanediol.

The possibility to achieve chiral separations with CPIL3 was tested by means of two different

chiral molecules: carvone and 1,2-transhexanediol. However, on the measurement of a

racemic solution containing these compounds, no peak broadening was observed.

Figure 6.7: Chromatogram of attempt at chiral separation on CPIL2 (5m x 0.25mm x 0.35µm) of a racemic

mixture of carvone (A) and Trans-1,2-cyclohexanediol (B)

However, in the chromatogram (Figure 6.7), only two peaks were observed, thus no chiral

separation was achieved. Upon further investigation on the peak widths, there also no

noticeable peak broadening occurring when comparing the enantiomerically pure sample

with the racemic mixture.

A

B

56

57

Chapter 7

Summary and conclusions

In this thesis, three chiral polymeric ionic liquid (CPIL) stationary phases were synthesized

and coated onto columns by means of the static coating method. Two of these stationary

phases, CPIL1 and CPIL2, were polymerised by means of chain-growth, the other, CPIL3, was

obtained by means of a step-growth polymerisation.

These stationary phases were tested on their thermal stability by means of

thermogravimetric analysis (TGA), which clearly showed that upon exchanging the halide

anion by a more thermally stable anion such as the bis(trifluoromethane)sulfonimide anion,

the thermal stability of the CPIL increased by approximately 50°C. This was made clear by

comparison of the onset degradation temperatures of CPIL1 and CPIL2 which were

approximately 250°C and 300°C respectively. The TGA measurements also made clear that in

terms of thermal stability, there was no significant difference between the chain-grown and

step-grown polymers as CPIL2 and CPIL3 both had the same onset degradation temperature.

It was also noticed that the maximum operational temperature (MaOT) of these stationary

phases when applied in gas chromatography were lower than the onset degradation

temperatures obtained by means of TGA, as by means of bleed measurements the MaOT of

CPIL1 and CPIL2 were determined to be around 200°C and 250°C respectively.

In terms of separation and retention, it was observed that poor peak shapes were obtained

for apolar compounds such as alkanes, whereas good peak shapes were obtained in the

separation of polar compounds. Although, the peaks obtained in the chromatogram were

rather broad. Chirality was tested by means of two chiral compounds, carvone and trans-1,2-

cyclohexanediol, but none of the synthesized stationary phases was capable of chiral

separation. Which is likely due to the limited size of the chiral methyl group on these

polymers.

It was observed that the CPIL2 stationary phase only yielded a limited amount of plates, as

the minimal plate height measured by means of Van Deemter curves was 0.99mm for a

thickness of coating of 0.35µm.

58

To broaden the scope of this thesis, the applicability of these CPIL stationary phases in

comprehensive GC (GCxGC) was assessed, where promising results were obtained, as

separation in the second dimension was achieved on a probe mixture containing multiple

different compounds.

The conclusion can be made that the attempt to achieve chiral separations with new

polymeric ionic liquid stationary phases was unsuccessful, however three new, never

synthesized high temperature stationary phases were obtained showing interesting

properties and possible applications in both GC-FID and GCxGC-FID, however further

research is required for further optimisation of these stationary phases.

Further research will be required to assess the applicability and plate heights of the

polymeric ionic liquid obtained by means of chain-growth polymerisation, CPIL3, as due to

time limitations the specifications (MaOT, plate height, …) of this stationary phase could not

be determined. More in-depth research into the influence of impurities on the quality of

coating, retention characteristics, and plate heights of columns coated with these

imidazolium-based ionic liquid stationary phases would also be interesting.

For the stationary phases synthesized in this master dissertation, it might also prove

interesting to see if derivatisation of chiral compounds would allow for chiral separation

since the time-limitations of this work didn’t allow for a thorough investigation thereof.

Appendix

A Reagents and materials

Imidazole, lithium hydride, bis(trifluoromethane)sulfonimide lithium salt, 1-vinylimidazole,

2,2’-azo-bis(isobutyronitrile) (AIBN), dodecane, pentadecane, aniline, benzaldehyde,

acetophenone, thionylchloride (SOCL2), methylsalycylate, (+)- and (-)-carvone, (R),(R)- and

(S),(S)-1,2-cycloexanediol, 3-nitrotoluene, dichloromethane (DCM, >99.8%), DMA, methanol

(MeOH, >99.9%), cinnamaldehyde, nonanoic lactone, propiophenone, butyrophenone,

hexanophenone, benzyl benzoate, toluene, anisole, sodium hydroxide pellets, hydrochloric

acid 37%, naphthalene, anthracene, THF (distilled in-house over sodium), acetone,

methanol, ethyl acetate, DMF (dried in-house over molecular sieves) and bare non-

deactivated fused silica with an internal diameter of 0.250 mm were obtained from Sigma-

Aldrich (Bornem, Belgium). R-(-)-bromo-2-methyl-1-propanol and (S)-(+)-1-bromo-2-

methylbutanewere obtained from ABCR GmbH (Karlsruhe, Germany).

Water (18.2 MΩ/cm) was purified and deionized in house via a Milli-Q plus instrument from

Millipore (Bedford, New Hampshire, USA). Deuterated solvents for all NMR spectroscopy

measurements: chloroform-d (CDCL3, >99.8%); DMSO-D6 (>99.9%), acetone-d (>99.9%), and

D2O (>99.9) were all purchased from Euriso-top.

Stock solutions of all measured compounds were prepared at 10mg/ml in dichoromethane

and diluted herein to 100µg/ml for all analyses.

B Equipment

B.1 NMR spectroscopy

A Bruker Avance 300 MHz Ultrashield and Bruker Avance II 400 MHz were used for the

measurement of all NMR spectra at room temperature.

B.2 Gas chromatography – Flame ionization detector (GC-FID)

All GC-FID measurements were performed on an Agilent 7890A system equipped with a

VWR Carrier-160 hydrogen generator. The selectivity studies were all performed with inlet

and outlet temperatures set at 200°C. Hydrogen was used as a carrier gas and mobile phase

velocities were 1.2mL/min for the selectivity studies and 1mL/min for the bleed

measurements. The Van Deemter measurements were performed isothermally at 130°C,

with an applied split-ratio of 10:1, whereas for selectivity studies a split of 2:1 was used.

B.3 Comprehensive GCxGC-FID

The comprehensive GCxGC measurements were performed on an Agilent 7890A with an FID

detector equipped with a VWR Carrier-160 hydrogen generator. Inlet and detector

temperatures were set at 250°C. Hydrogen was used as a carrier gas and mobile phase

velocities were set at 0.6mL/min for the first dimension and 30mL/min for the second

dimension. Modulation was set at 2.3sec with a sample time of 1.5sec and a temperature

program was set from 50°C to 250°C at 5°C/min. The 2D measurements were performed by

means of splitless injection.

C NMR-spectra

C.1 Monomer CPIL2

Figure C.1: 1H-NMR spectrum of CPIL2 monomer

A

J

B C

D E F

G

H I

A

A H

G D C

B

E+F

J

I

C.2 CPIL2

Figure C.2: 1H-NMR spectrum of CPIL2

C.2 Synthesis CPIL3

Figure C.2.1: 1H-NMR spectrum of CPIL3 monomer intermediate

A

B E

F G

H C D

A

B

C D

E

F

G H

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Development Of Chiral And Achiral Polymerized Ionic Liquid Stationary Phases

For High Temperature Gas Chromatography

A. Ampe; F. Lynena

a Separation Science Group, Department of Organic and Macromolecular Chemistry,

Ghent University, Krijgslaan 281 S4-Bis, 9000 Ghent, Belgium

In this study the possibility to synthesize chiral polymerized

imidazolium based ionic liquid (CPIL) stationary phases for

applications in high temperature gas chromatographic is

investigated. Two high temperature stable stationary phases were

synthesized through a chain-growth polymerization step. These

two synthesized CPILs consist of the same cation but have

different anions. The thermal stabilities were assessed using

thermal gravimetric analysis (TGA) and was compared with

bleeding profiles obtained from the statically coated GC columns

(5m x 0.25mm x 0.35µm). The CPIL containing a halide anion,

CPIL1, showed an initial degradation at around 250°C, whereas

CPIL2, consisting of a bis(trifluoromethane)sulfonimide showed

an initial degradation of 300°C. The CPIL2 stationary phase was

further investigated upon and minimal plate heights of 1mm –

1.5mm were obtained for retained solutes such as nonanoic lactone

and benzyl benzoate. Even though these stationary phases showed

promising thermal stabilities, no chiral selectivity was observed.

Keywords: Gas chromatography, Polymerised ionic liquid phase, Imidazolium,

Chiral separations, Chiral stationary phases

1. Introduction

Even though high-performance liquid chromatography (HPLC) is inherently more

limited in terms of achievable efficiency and resolution compared to what can be

achieved by gas chromatography (GC), chiral separations are nowadays still usually

performed by HPLC instead of GC. This is partly due to the limited maximum

operational temperatures (MaOT) of only 230°C for commercially available chiral

stationary phases for gas chromatography.

Therefore, improvement towards more efficient (chiral) GC columns capable of better

separations is more than relevant. Since an increase in thermal limits in GC would lead to

greener and more efficient ways to analyze these kinds of solutions.

In the long history of separation science, the discovery of ionic liquids (ILs) and

application thereof as stationary phases in GC and as stationary and/or mobile phases in

LC has allowed for significant breakthroughs regarding e.g. the increase of thermal limits

of the ensuing chromatographic methods.

These ionic liquids are considered new types of molecules and generally consist of an

organic cation paired with an organic or inorganic anion. (9) The main reason for the

sudden increase in research towards ionic liquids in separation sciences, is due to several

interesting properties of this kind of molecules. The most important ones being their high

thermal stability, their high polarity, negligible vapour pressure, and their dual nature

which allows them to effectively separate both polar and apolar compounds as if they

were a polar or apolar stationary phase respectively. (8)

It is mainly due to their high thermal stability that these types of molecules have

become an interesting candidate for synthesis of chiral stationary phases (CSP) with

higher maximum operational temperatures than commercially available stationary phases.

There are generally two ways to generate a chiral stationary phase with ionic liquids:

either a certain amount of chiral selector can be added to an achiral ionic liquid, or the

ionic liquid itself can be made chiral. (7) Most research so far has been performed on

adding permethylated cyclodextrins (CD) to achiral ionic liquids to achieve high

temperature stable chiral stationary phases. (7) However, the MaOT temperatures of such

stationary phases were generally as limited as the commercially available CSPs.

Therefore, in this study an attempt will be made to create an ionic liquid chiral

stationary phase in which the cation itself is chiral. Since it was shown in literature that

the thermal stability of ionic liquids increases when they are polymerized (3), the cation

will be made in such a way to allow polymerization to further increase the thermal

stability.

2. Experimental procedure

2.1 Reagents and materials

Imidazole, lithium hydride, bis(trifluoromethane)sulfonimide lithium salt, 1-

vinylimidazole, 2,2’-azo-bis(isobutyronitrile) (AIBN), dodecane, pentadecane, aniline,

benzaldehyde, acetophenone, thionylchloride (SOCL2), methylsalycylate, (+)- and (-)-

carvone, (R),(R)- and (S),(S)-1,2-cycloexanediol, 3-nitrotoluene, dichloromethane (DCM,

>99.8%), DMA, methanol (MeOH, >99.9%), cinnamaldehyde, nonanoic lactone,

propiophenone, butyrophenone, hexanophenone, benzyl benzoate, toluene, anisole,

sodium hydroxide pellets, hydrochloric acid 37%, naphthalene, anthracene, THF

(distilled in-house over sodium), acetone, methanol, ethyl acetate, DMF (dried in-house

over molecular sieves) and bare non-deactivated fused silica with an internal diameter of

0.250 mm were obtained from Sigma-Aldrich (Bornem, Belgium). R-(-)-bromo-2-

methyl-1-propanol and (S)-(+)-1-bromo-2-methylbutanewere obtained from ABCR

GmbH (Karlsruhe, Germany).

Water (18.2 MΩ/cm) was purified and deionized in house via a Milli-Q plus

instrument from Millipore (Bedford, New Hampshire, USA). Deuterated solvents for all

NMR spectroscopy measurements: chloroform-d (CDCL3, >99.8%); DMSO-D6

(>99.9%), acetone-d (>99.9%), and D2O (>99.9) were all purchased from Euriso-top.

Stock solutions of all measured compounds were prepared at 10mg/ml in

dichoromethane and diluted herein to 100µg/ml for all analyses.

2.2 Synthesis of CPIL obtained using a chain-growth mechanism

The other CPIL was synthesized via a chain-growth mechanism as shown in Figure 2,

based on the same reaction performed in the work of Marcilla et al. (1) and Green et al.

(2) The ionic liquid monomer was synthesized by adding 8mL of (S)-(+)-1-bromo-2-

methylbutane to a solution containing 5mL 1-vinylimidazole in 20mL dry ethyl acetate.

The reaction was then heated to 77°C to reflux and left to react for 48h. During the

reaction, a separate viscous layer formed at the bottom of the flask. After decanting and

washing with ethyl acetate, the product was dried using a rotary evaporator to remove

residual reagents and solvent.

Next, 4.040g of the monomer was dissolved into 15mL dry DMF and 2mol% of AIBN

(0.33mmol) was added as initiator. The solution was then purged with argon to remove

traces of oxygen. The mixture was the heated to 70°C and left to stir for 12h. The

polymer was then precipitated as a white powder by adding an excess of acetone to the

mixture. This powder was filtered off over a sintered glass filter and washed with acetone.

The resulting precipitate was collected and dried in a vacuum oven. This polymer was

named CPIL1.

To increase the thermal stability, the bromine anion was exchanged for the more

thermally stable bis(trifluoromethane)sulfonimide anion. This was performed by

dissolving 2g of the CPIL1 polymer in 7mL of water, adding a 3-equivalent excess of

bis(trifluoromethane)sulfonimide lithium salt (6.959g) in 7mL water, and stirring for 24h.

The resulting precipitation was filtered and rinsed with water, until no more chloride was

detected in the rinsing water using AgNO3. The resulting product was named CPIL2.

Figure 1. Synthesis of the chain-growth monomer followed by polymerization towards

CPIL1 and ion exchange to obtain the CPIL2 polymeric ionic liquid.

2.3 Characterisation of monomers and polymers

NMR data of monomers and polymers were obtained on a Bruker 400MHz

UltrashieldTM and Bruker 300MHz UltrashieldTM and are provided in the supplementary

information. Thermal gravimetric analyses were performed on a TGA system in a

temperature range from 25°C to 800°C at 10°C/min under ambient air.

2.4 GC capillary column coating

Both polymers were coated onto a capillary column using the static coating method.

(4) Therefore, CPIL1 was dissolved into dichloromethane, and CPIL2 was dissolved into

acetone. The desired film thickness is dependent on the applied concentration and can be

calculated using [1] with c = concentration of the polymer solution (in mg/ml), df =

desired film thickness, ρ = density of the polymer, and dc = internal diameter of the

column. The densities of the polymers were estimated to be 0.7019 g/ml based off the

work of Roeleveld et al. (3)

Therefor the polymer solutions were made with a concentration of 4mg/ml to obtain a

film thickness of 0.35µm onto the capillary column with internal diameter of 0.25mm.

c = 4df*ρ/dc [1]

The columns were pretreated as followed: first a NaOH solution of 1M was put into a

pressurized glass-container and nitrogen was applied to flush the column, after which the

column was rinsed with milli-Q water. Next up, a 1M HCl solution was sent through the

column after which it was rinsed again with milli-Q water. Then, MeOH was flushed

through to remove traces of water. The column was then flushed with the solvent of the

polymer solution, after which the polymer solution was sent through the column. When

the capillary was filled with polymer solution, the nitrogen pressure was lowered to

atmospheric pressure and the end of the end of the column was submerged in silicone for

sealing. The column was left like this for 30min to obtain proper sealing, afterwards the

other end of the column was attached to a vacuum pump and the column was put into a

temperature-controlled water bath set at 25°C for CPIL1, and 40°C for CPIL2 for

evaporation of the solvent, leaving a thin layer of polymer on the inner walls of the

capillary. After coating, the columns were conditioned for two hours at 120°C in a GC-

FID under a hydrogen flow of 1.2mL/min to increase the longevity of the stationary

phase.

2.5 Chromatographic conditions for GC

All GC-FID analyses were performed on an Agilent 7890A system equipped with a

VWR Carrier-160 hydrogen generator. The inlet and outlet temperatures were set at

200°C to prevent thermal degradation of the coating. Hydrogen was used as a carrier gas

and mobile phase velocities were 1.2mL/min for the selectivity studies and 1mL/min for

the bleed measurements. The temperature program used for the bleed measurements

started at a temperature of 40°C up to the maximum applied temperature at a rate of

10°C/min, after which the maximal temperature was held for another 10min. For the

selectivity studies, a split ratio of 2:1 was applied in all measurements. The Van Deemter

measurements were performed isothermally at 130°C, with an applied split-ratio of 10:1.

In all experiments the FID was operated at 225°C, with following gas flow settings:

H2 = 30mL/min, Air = 300mL/min, N2 = 30mL/min.

3. Results and discussion

3.1 Thermal properties of the CPIL columns

Figure 2. Thermal gravimetric analysis for the CPIL1 and CPIL2 stationary phases.

Temperature program: 25°C – 800°C at a rate of 10°C/min under air flow

The thermal stabilities of CPIL1 and CPIL2 were assessed by means of TGA analysis.

As can be observed in the thermogram for CPIL1 and CPIL2 (Figure 2), an initial weight

loss was detected at around 100°C for CPIL1, which is related to the hygroscopic nature

of this polymer. (1) Upon exchanging the chloride anion with

bis(trifluoromethane)sulfonimide, an increase of degradation temperature is detected of

around 50°C, confirming the thermally stabilizing effect of this anion. The onset

degradation temperatures were found to be approximately 250°C for CPIL1 and 300°C

for CPIL2.

To fully assess the applicability of this kind of polymers as stationary phases, TGA

measurements prove incomplete since at lower temperatures, degradation can already

occur which would cause disruptions in the homogeneity of the stationary phase.

Therefor, CPIL1 and CPIL2 were also tested after they were coated onto columns with

dimensions of 5m x 0.25mm i.d. x 0.35µm by means of the static coating method. (4)

These bleed results (Figure 3) clearly indicated that the onset degradation temperature

obtained by TGA measurements is an over-estimation of the applicability by about 50°C,

since the background in these measurements already reached the threshold value of 20pA

at around 200°C for CPIL1 and 250°C for CPIL2. Since bleeds above the threshold of

20pA indicate significant degradation in terms of lifetime of the column, the maximum

operational temperatures for CPIL1 and CPIL2 were thus determined to be 200°C and

250°C respectively.

Figure 3. Bleeding profiles of prepared CPIL1 and CPIL2 columns for a blank analysis

on FC-FID with a temperature program starting at 40°C and ending at different maximum

temperatures held for 10min.

3.2 Column efficiency study

Due to CPIL1 and CPIL2 consisting of the same cation and the assumption was made

that for column efficiency and separations, the influence of the anion is negligible enough.

To determine the optimal operational conditions of the columns and the efficiency thereof,

Van Deemter curves were constructed via isothermal GC-FID experiments for two probe

compounds: nonanoic lactone and benzyl benzoate. The plots for these compounds are

depicted in Figure 4.

It was found that the optimal mobile phase velocities on the synthesized stationary

phases were between 25 and 35cm/s where minimal plate heights of 1 – 1.5mm were

obtained, which corresponds to around 1000 plates/m for the columns made in this work.

This value is rather low compared to e.g. the amount of plates obtained in the work of

Roeleveld et al. (3), who managed to obtain 2000 – 2500 plates/m with similar polymeric

ionic liquid stationary phases. The reason thereof is likely due to either an increase of

impurities which were present in the polymeric ionic liquids synthesized in this work or

could be an influence due to the extra methyl group present on the cation moiety.

Additionally, assessment on separation characteristics of the stationary phases was

performed with a 30m column coated with CPIL2. For this, two different mixtures were

injected onto the column (30m x 0.25mm x 0.35µm) to determine whether separations

could be achieved. The chromatograms of these measurements can be found in Figure 5.

For the mixtures applied, separation was achieved with good peak shapes, however it

can be observed that the peaks in the chromatogram are rather broad, which is likely due

to the low column plate numbers of only 1000 plates/m, as mentioned above.

Figure 4. Constructed Van Deemter curves of nonanoic lactone and benzyl benzoate

measured on a column coated with CPIL2. (5m x 0.25mm x 0.35µm)

Oven temperature: 130°C.

Figure 5. Analysis of a sample containing 100ppm of benzaldehyde (A),

methylsalycylate (B), 3-nitrotoluene (C), ɣ-nonanoic lactone (D), and benzyl benzoate

(E) measured on a column coated with CPIL2 (30m x 0.25mm x 0.35µm)

3.3 Assessment of chiral separations

One of the main goals of this work, was to synthesize imidazolium-based polymeric

ionic liquids which were capable of chiral separations. However, no peak broadening was

observed in the chromatograms of the measurement of two chiral compounds: carvone,

and 1,2-trans-cyclohexanediol. (Figure 6) The lack of any chiral recognition was likely

due to either racemization of the stationary phase during any of the synthesis steps, or the

fact that a single methyl group as chiral selector is too insignificant to achieve chiral

separations. However, further research will be required e.g. derivatization of chiral

A B

C

D

E

compounds and/or more tests on different chiral molecules, to determine the cause of lack

of chiral recognition.

Figure 6. Chromatograms for the assessment of chiral separations on a CPIL2 column

(5m x 0.25mm x 0.35µm) of a racemic mixture of carvone (A) and trans-1,2-

cyclohexanediol (B).

4. Conclusion

In this study two (chiral) polymeric ionic liquid (CPIL) stationary phases were

synthesized and coated onto columns by means of the static coating method. These

stationary phases were tested on their thermal stability by means of thermo gravimetric

analysis (TGA), which showed that upon exchanging the halide anion of an ionic liquid

by a bis(trifluoromethane)sulfonimide anion, the thermal stability increases by

approximately 50°C. The maximum operational temperatures of the synthesized

imidazolium-based ionic liquids were around 200°C and 250°C for CPIL1 and CPIL2

respectively.

In terms of separation of compounds, it was observed that analysis of apolar

compounds such as alkanes resulted in bad peak shapes, whereas polar to highly polar

compounds resulted in good peak shapes. The minimal plate height for CPIL2 were

determined by measuring Van Deemter plots, which showed that for retained compounds

such as nonanoic lactone and benzyl benzoate the minimal obtained plate heights were

around 1mm.

The ability to achieve chiral separations was assessed for the CPIL2 stationary phase,

but it was found that for the two probe compounds: carvone and trans-1,2-

cyclohexanediol, no chiral separation was achieved. There are three plausible causes why

chiral separations weren’t achieved: radical polymerization led to racemization of the

chiral center, a single methyl group doesn’t suffice to achieve chiral separations.

Although further research is required to asses if the lack of chiral selectivity is due to one

of the possibilities above or due to another reason.

Acknowledgments

Adriaan Ampe gratefully acknowledges the assistance of Mathijs Baert for helping

with the synthesis, both by assisting with the experimental work as critical discussions for

progress in the synthesis.

A B

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