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Know the Difference
When to use guard columns and retention gaps in GC
Jaap de Zeeuw
Figure 1: Principle of the retention gap
Click to enlarge. |
Guard columns and retention gaps are widely used in gas chromatography. However, many users have difficulty understanding the difference between these two products, and there is, in fact, a significant difference in applications. Retention gaps are mainly used for focusing the sample components when introducing a large liquid sample directly on to the column. The guard column, conversely, is used to protect any analytical column for contamination. When using a retention gap system, the gap will also act as a guard column, but its primary function is to create a focusing effect.
Such guard columns and retention gaps must be coupled, which traditionally introduces a potential risk in many labs. But a new approach has been launched where the guard column is integrated with the analytical column. By applying a “segment”-coating technology, the stationary phase can be deposited in a certain part of the column, allowing for a deactivated section at the beginning. Column coupling is no longer required and maintenance is greatly simplified.
Use of retention gaps
In today’s laboratory, GC methods must be simplified and faster, and low detection limits are requested, not to mention that sufficient precision must also be obtained. It all starts by introducing the sample in the smallest possible injection band and to make that band migrate through the capillary with minimal loss of the components being analyzed.
With an on-column injection, a liquid sample is directly introduced into the capillary column as a liquid, while the capillary column is kept at a temperature of 10 to 15°C below the solvent boiling point. During this process, the sample components are spread in a non-reproducible way over the first 20 to 100 cm of capillary while the solvent is evaporating. Parameters such as injection speed, carrier gas flow, solvent and column temperature, type of solvent, and pressure will all affect the injection band width.
Figure.2: Segment coating. (A) Fill complete column, push plug through to Position A and start coating process. (B) When coat front has reached Postion B, residue is pushed out. |
Additionally, when non-bonded stationary phases are used, any direct contact with liquids will result in a distortion of the stationary phase film and short lifetime of the capillary. The majority of today’s stationary phases, such as Rtx and Rxi, are immobilized by cross- and surface-bonding techniques.
For proper application of the on-column injection technique, the use of retention gaps is essential (Figure 1). The retention gap consists of a length of 1 to 3 m of deactivated capillary that is positioned in front of the analytical column. All the processes described will still take place, but now the components are distributed over the retention gap.
Once the oven temperature is increased, the sample components will start to move (note that there is very little retention; thus it’s called a retention “gap”). When reaching the analytical column, the components will focus in the stationary phase resulting in a narrowing of the injection band width.
As these retention gaps are mainly used for on-column injection, the inside diameter usually measures 0.32 to 0.53 mm, and the needle of an on-column syringe must be able to enter the retention gap. For coupling of the retention gaps with the analytical column, we generally need devices that can deal with different diameters of capillary tubing.
Retention gaps and splitless injection
For splitless injection, we generally do not require a retention gap. The sample is injected in a hot injection port, evaporates, and then is transported with the carrier gas flow of approximately
1 mL/min in to the capillary. The amount of solvent vapor that enters the column per time unit is much smaller than with on-column injection. Although with splitless injection the oven temperature is also 10 to 15°C below the solvent boiling point, there will be little chance that solvent condensation takes place. The high concentration of solvent entering the capillary will cause a strong focusing effect for the components, generating a narrow injection band. If, in splitless injection, a method is used where the initial (injection) oven temperature is much lower than the boiling point of the solvent, the risk of solvent condensation (forming a liquid plug) will increase, which can cause unwanted broadening of the injection band. Coupling a retention gap will also “fix” this problem.
Wettability of retention gap
Figure 3: New AlumaSeal connections: leak-tight coupling of fused silica of different diameters, as well as MXT capillary tubing |
Important for a well-performing retention gap is the fact that the retention gap surface must be “wettable” for the solvent used. The solvent also needs to spread evenly over the surface, meaning that non-polar solvents (hexane, methylene chloride, isooctane, benzene) require non-intermediate deactivated retention gaps, and the more polar solvents (methanol) will require polar-deactivated retention gaps. If the polarity of retention gap and solvent do not match, the solvent will form droplets inside the capillary. The carrier gas will then “push” this droplet along the retention gap into the analytical column, resulting in a broadened injection and even peak splitting.
Retention gaps for large-volume injection
Rather than the injection of 1 mL on a 1- to 2-m retention gap, chromatographers can also inject much larger amounts on much longer retention gaps. Here, we talk about large-volume injection techniques where retention gaps of 8 to10 m in length are used. Such retention gaps can be loaded with 100 to 200 mL of sample. Injection must be slow to allow the solvent to evaporate while passing through the retention gap.
With large-volume injection, detection limits can be reduced by a factor of 100. The technique requires some skills to optimize all injection parameters. Additionally, the large-volume retention gaps pollute relatively quickly due to the large amounts of sample introduced.
Use of guard columns
As mentioned earlier, often the guard column is confused with the retention gap. A guard column can be put in front of any analytical column, and the main application protects the analytical column from contamination. We need to use a retention gap when we introduce a liquid sample onto the column that is causing the poor injection. The retention gap “fixes“ that, and using guard columns will allow more analysis to be done before maintenance.
Figure 4: Integra-Guard can be recognized by a separate string. |
Guard columns are widely used in GC and LC separation technology to protect the analytical column from contamination .The sample that is introduced in the column is not always pure. Although the best chromatography is obtained with “clean” samples, the practical situation is that sample clean-up procedures are minimized, and that relatively “dirty” samples are fed into the column. Samples can contain particulates, heavy components, derivatization reagents, iogenic residues, acids and/or bases. All of these compounds can interfere with the stationary phase and they will influence the separation process. Usually the degradation of column performance is a slow process, but it will happen. Most of the time, these impurities accumulate in the first meter(s) of the column, and, by cutting off this section, the separation is restored.
Many users choose to connect a guard column in front of the analytical column. Such a guard column is deactivated and can be trimmed when polluted and then eventually replaced. The type of guard column can be chosen, whether by length, internal diameter and the type of deactivation. One has different choices of guard columns, which may consist only of deactivated capillary, or it can be a coated capillary. Also, depending on the application guard columns have a lifetime of one week up to six months.
Deactivated capillary tubing: Deactivated fused silica tubing can be purchased per meter, and a defined length is coupled in front of the analytical column. Upon contamination, a section of the guard column is removed. When the whole guard is “consumed,” a new guard column can be coupled. A disadvantage of cutting parts of the guard column is that the column becomes shorter, resulting in changed retention times. However, if a similar length is always cut from the guard column, the change in retention time becomes very predictable.
A deactivated guard column will also result in band focusing. If the injection is not optimal, there will be a focusing effect similar to that of the retention gap.
Figure 5: Design of Integra-Gap with MXT-type columns. Click to enlarge. |
Coated capillary tubing as guard: A different way of using guard columns is to use one that is coated. As the guard column needs to prevent contamination of the analytical column, a coated guard column can help, as it has, in addition to surface deactivation, a stationary phase layer. And using a coated guard column produces no focusing effects.
The easiest and most economical way for using coated guard columns (or pre-columns) is to buy not one, but two analytical columns. We will use one as a separation column, and the other as a coated guard column. From this second column, we will cut 2-m sections and couple a section in front of the analytical separation column. We will run our samples until contamination affects our peak shape/response, and then we can replace the guard for a new 2-m section.
The system we have created will reproduce retention times, since we will always replace the entire 2-m coated guard column. Since the stationary phase is the same on the guard as it is on the analytical column, there will be no surprises. The coated guard column will also allow more aggressive samples and more contamination before it gives up. Lastly, we are able to cut 15 coated guard columns from a full 30-m analytical column…very economical!!
Coupling guard columns/retention gaps
The coupling between capillary tubing can be done by several different devices. Press-tight connectors are very popular, but practically speaking, users experience problems making a leak-tight coupling. It’s difficult to get a good seal; when running MS-methods, the mass spectrometer will often detect the presence of a leak. Also, the fused silica sometimes slips out. However, VU2 unions already show improved stability due to the extra fixation of the fused silica.
Connection devices that contain ferrules work much better, although you sometimes need three hands to make the coupling. Vespel-polyimide ferrules also have to be re-tightened once they have been heated. Most of these coupling devices have relatively high thermal capacity…they need a body to keep the ferrule in place.
One of the latest developments in column coupling is the AlumaSeal. This connection is leak tight from sub-ambient up to temperatures of 350°C and can be used with challenging applications such as vacuum GC and GC-MS. The seal allows for the coupling of metal and fused silica with different diameters.
Integra-Guard or Integra-Gap by applying segment coating
Figure 6: Analysis of biodiesel using MXT-Biodiesel TG after 1 and 100 cycles to 430°CClick to enlarge. |
Restek has a long history in coating capillary columns. As there are different processes possible for making GC columns, one of the widely used methods found in literature is the static coating process. Here, the capillary column is filled with a coating solution of stationary phase in a volatile solvent. The column is sealed on one end and, on the other side, a vacuum is applied. The solvent is evaporated and then the dissolved polymer is deposited on the deactivated inside wall of the fused silica column.
The static coating method also allows for coating columns by segment. Figure 3 shows a schematic setup of such a process. When filling, for example, a 40-m capillary with the coating solution, only the last 30 m are filled. When applying vacuum, the deposition of stationary phase starts at point A. The first part remains uncoated, having only the deactivation (which is deposited there in a previous procedure).
Once the coating process is started, and depending on column length, diameter, type of solvent and temperature, point B is reached in a certain time. When it is reached, the vacuum is stopped and the coating solution is flushed out of the last coils by a flow of nitrogen. In this way it is possible to deposit the stationary phase in a designated portion of the capillary, creating the Integra-Guard or the Integra-Gap. Similar technology can also be used to make integrated transfer lines. One can also fill the column completely and use pure solvents in the section that needs no deposition of a stationary phase.
The advantages are clear. As there is no coupling present, we have all the benefits of having “no coupling:”
• There is no connection to make, thus saving time
• No leaks improves stability and provides more accurate data
• No dead volumes with activity or thermal mass
• Maintenance is easy, and the solution is integrated.
Integra-Guard columns are 5 to 10 m in length and can be made for most standard stationary phases and all column diameters. What’s important here is to make a clear mark where the guard column starts, done by a separate high-temperature string or by a different label (Figure 4).
Also, Integra-Gap technology is possible, which is mainly applied for larger-diameter fused silica or MXT capillaries. As for on-column injection, a wide-bore column is required because of the syringe needle’s outside diameter. A 0.53-mm capillary must be used as a retention gap and also for the separation capillary. Figure 5 shows an example of an MXT 0.53-mm column, which was developed specifically for biodiesel analysis. In this application, trace triglycerides need to be measured and an on-column technique is recommended. To elute the triglycerides, the column must be heated up to 380 to 4 00°C, which limits the use of fused silica. In addition, column connections are critical because of dead volumes, thermal mass, activity and possible leaks.
The Integra-Gap, along with the MXT-Biodiesel TG columns, allows a simple solution for detecting trace biodiesel without hassling with retention gaps, cutting and coupling. Figure 6 shows an analysis of biodiesel on a 16-m 3 0.53-mm MXT Biodiesel TG with a 2-m Integra-Gap, using on-column injection. Note the perfect peak shape for the solvent. A temperature program was used up to 430°C. After 100 cycles, there was no significant change in retention or separation efficiency. Also ,Triglyceride response was very good. Such solutions will be welcome in the routine laboratory as column coupling can be minimized.
Integrated guard and gap solutions will contaminate, and periodically, a piece must be cut. When the whole guard or gap is consumed, you still can make the decision to couple or to use a new integrated solution. Whatever the choice is, you have at least experienced the possibility to reduce or even eliminate the column coupling.
Jaap de Zeeuw is the International
GC Consumables Specialist at
Restek Corp. He may be contacted at ChromatographyTechniques@advantagemedia.com.
Restek Corporation
110 Benner Circle
Bellefonte, PA, 16823
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