I. Instrument Preparation
Gas Purification
Make sure the carrier gas is clean. The carrier gas MUST contain less than 1ppm of oxygen, moisture, or any other trace contaminants to prevent column degradation, decreased column lifetime, and increased stationary phase bleed. Contaminants such as trace hydrocarbons cause ghost peaks to appear during temperature programming and compromise the validity of analytical data. Make-up gas should also be contaminant-free or baseline fluctuations and excessive detector noise can occur. Detector gases, such as hydrogen and compressed air, should be free of water and hydrocarbons or excessive baseline noise can result. We highly recommend installing a High Capacity Indicating Oxygen Trap on both the carrier and make-up gas lines. Because moisture reacts with most oxygen traps, we suggest installing a Moisture Trap in front of the oxygen trap. (Install purifiers as closely as possible to the GC's bulkhead fitting, not system-wide. If purifiers are installed system-wide, a leaky fitting downstream of the trap could allow oxygen and moisture to enter the gas stream and degrade column performance.) The Moisture Trap can also be used on the FID air line or the ECD make-up gas line to eliminate noisy, rolling baselines when operating at high detector sensitivities. If hydrocarbon contamination is suspected, install a Hydrocarbon Trap between the moisture and oxygen traps. To prevent spontaneous breakage, coil the line leading to and from the purifiers to relieve strain and isolate instrument vibrations. | A note on equipment: Because oxygen, moisture, and elastomeric contaminants can migrate through rubber or elastomeric diaphragms and enter the carrier gas, all regulators should be equipped with stainless steel diaphragms. |  | Traps shown:
A. Moisture Trap:
Super-Clean™ Ultra-High Capacity Moisture Filter (cat.# 22028) B. Hydrocarbon Trap:
Super-Clean™ Ultra-High Capacity Hydrocarbon Filter (cat.# 22030) C. High Capacity Indicating Oxygen Trap:
Super-Clean™ Ultra-High Capacity Oxygen Filter (cat.# 22029) | Carrier Gas Selection A fast carrier gas that exhibits a flat van Deemter profile is essential in obtaining optimum capillary column performance. Because capillary columns average over 30 meters in length (compared to 2 meters for packed columns), a carrier gas that minimizes the effect of dead time is important. In addition, because capillary columns are head pressure controlled, not flow-controlled like most packed columns, the carrier gas flow decreases by 40 percent when programming from ambient to 300°C. Therefore, a carrier gas that retains high efficiency over a wide range of flow rates is essential for obtaining good resolution throughout a temperature-programmed analysis. Figure A shows the van Deemter profile for hydrogen, helium, and nitrogen carrier gases. The curves were generated by plotting the Height Equivalent to a Theoretical Plate (H.E.T.P., the length of the column divided by the total number of theoretical plates) against the column's average linear velocity. The lowest point on the curve indicates the carrier gas velocity at which the highest column efficiency is reached.
Hydrogen is the fastest carrier gas (uopt: 40cm/sec.) and exhibits the flattest van Deemter profile. Helium is the next best choice (uopt: 20cm/sec.). The head pressures at optimum flow rates are similar for hydrogen and helium because hydrogen has half the viscosity and double the linear velocity of helium. Nitrogen's performance is inferior for capillary columns and is usually not recommended because of the slow optimum linear velocity (uopt: 12cm/sec.) and steep van Deemter profile. Figure B compares hydrogen to helium in an isothermal analysis using optimum linear velocities. The resolution is similar, but the analysis time is reduced by 50 percent when using hydrogen. However, helium does improve the separation of very low boiling or early eluting compounds since they allow more interaction with the stationary phase. Temperature programming usually provides similar analysis times between hydrogen and helium since the elution of most compounds strongly depends on the oven temperature. Therefore, the savings in analysis times are not as noticeable as when using isothermal oven conditions. Figure C illustrates that hydrogen is only slightly faster than helium when both carrier gases are operated under the same temperature-programmed conditions. Also, note that helium improves the resolution of the early eluting compounds (peaks 1 & 2). Exert Caution when using Hydrogen as a Carrier Gas Hydrogen is explosive when concentrations exceed 4 percent in air and should only be used by individuals who have received proper training and understand the potential hazards. Proper safety precautions should be taken to prevent an explosion in the oven chamber. Some gas chromatographs are designed with spring loaded doors, perforated or corrugated metal oven chambers, and back pressure/flow controlled pneumatics which minimize the hazards when using hydrogen carrier gas. Additional precautions include: - Frequently checking for leaks using a thermal conductivity leak detector (cat.# 22839).
- Minimizing the amount of carrier gas that could be expelled in the oven chamber if a leak were to occur by installing a needle valve, restrictor, or flow controller prior to the carrier inlet bulkhead fitting (only necessary for head pressure controlled systems).
- Purging an inert gas (N2) into the oven chamber to displace oxygen and prevent an explosive atmosphere from forming.
Hydrogen is expelled from both the split vent and septum purge when it is used as a carrier gas. Because of hydrogen's fast diffusivity, an explosion in a laboratory setting is highly unlikely. However, a spark from static electricity can ignite the hydrogen exiting from a septum purge or split vent which could cause a flame. Precautions to minimize the problems with hydrogen exiting the split vent or septum purge include: - Plumbing the exit lines to a hood or venting the escaping gas outside.
- Plumbing the lines to exit into a vial of water.
- Plumbing the exit lines to a position where analysts could not get burned if inadvertent ignition occurred.
Flow Regulated/Back Pressure Pneumatic Systems
Figure D illustrates a flow regulated/back pressure system commonly used for split/splitless inlets (HP-5890, Varian 3300, 3500, & 3600 GCs). A flow controller positioned upstream of the injector controls the total amount of carrier gas that is expelled from the split vent, septum purge, and column (knob labeled "Total Flow" on HP 5890 GCs). The back pressure regulator (knob labeled "Column Head Pressure" on HP 5890 GCs) stops or reduces the flow from exiting the split vent until the desired column head pressure is reached. The flow controller provides the increase of pressure necessary to meet the requirements of the back pressure regulator. It is the back pressure regulator, located downstream of the split point, that actually controls the capillary column flow rate. One of the primary benefits of a flow controlled/back pressure regulated system is that adjustments to the capillary column flow rate (via head pressure changes) do not affect the amount of carrier gas exiting the splitter vent. Once the desired split vent flow rate is achieved, the flow controller setting should not have to be changed when installing different columns.
Flow regulated/back pressure systems prevent a drastic carrier gas loss that can happen if an inlet fitting or column leak occurs. Leaks are indicated by a failure to obtain the proper operating pressure with the capillary column. A common mistake is to increase the total system flow by turning the flow controller up when a proper head pressure cannot be obtained rather than checking for inlet leaks. Head Pressure Regulated Pneumatic Systems
Figure E illustrates a head pressure regulated inlet system used in some split/splitless inlet systems (PE Autosystem, 8500, and Shimadzu 9A and 14A GCs). A single-stage pressure regulator is used to control the flow rate in the capillary column by increasing or decreasing the upstream inlet pressure. The split vent and septum purge flow rates are controlled by a needle valve or variable restrictor located downstream of the pressure regulator. Head pressure systems require adjustment of the needle valve controlling the septum purge or split vent every time a change is made in the column's head pressure.
We recommend placing a throttling valve (needle valve or restrictor) on the carrier gas inlet bulkhead fitting of pressure-regulated systems to prevent a catastrophic carrier gas loss should an inlet leak occur. If several GCs are attached to a common carrier gas source, a leak in one GC could drain the carrier gas from all other GCs, causing a loss of flow and subsequent damage to all capillary columns in the entire system. To prevent this from happening, limit the flow of carrier gas to each gas chromatograph (via a throttling valve) until it matches the flow requirements of the inlet system. This can be detected when the column's head pressure starts to decrease if the throttling valve is closed any further. Injector Maintenance Perform injector maintenance prior to installing a capillary column. Periodic maintenance is required after installation, depending on the number of injections and the cleanliness of the samples. Maintenance includes cleaning and deactivating inlet sleeves, replacing critical inlet seals, and replacing the septum. Review the instrument manual inlet diagram prior to disassembly. Cleaning and Deactivating Injector Sleeves
Don't install a new Restek column with a dirty injector sleeve! For optimum column performance, the inlet sleeve needs to be free of septum particles, sample residue, and ferrule fragments. Use deactivated inlet sleeves when analyzing samples with active functional groups or compounds prone to decomposition or adsorption onto untreated glass surfaces.  If the sleeve is deactivated and not excessively dirty, cleaning with organic solvents will not affect the integrity of the deactivation layer. First, use non-swelling organic solvents such as methanol or isopropyl alcohol to remove septum particles that adhere to the sleeve wall. Next, use solvents such as pentane, methylene chloride, or toluene to remove sample residue. Nylon tube brushes and pipe cleaners (cat.# 20108) are ideal for cleaning sleeves. Do not use laboratory detergents, acids, or bases to clean sleeves because they will remove the deactivation layer and require sleeve resilanization. Sleeves that are very dirty or contain pyrolyzed residue can be difficult to clean. Heating borosilicate or quartz sleeves overnight in a muffle furnace at 550°C will remove most contaminants. Etching with a 1:1:1 mixture of hydrofluoric acid, sulfuric acid, and deionized water for ten seconds is also very effective. However, heating to 550°C or using the acid-etch procedure will remove the deactivation layer and require sleeve resilanization. Caution - Exert extreme caution when using hydrofluoric acid. Hydrofluoric acid can cause severe burns and nerve damage if it is ingested, inhaled, or brought into contact with the skin. Only properly trained professionals equipped with the appropriate safety devices should attempt to handle strong acids. Most new sleeves received from instrument manufacturers are not deactivated. Before optimum column performance can be achieved, sleeves must be deactivated. Restek has developed a unique, high temperature silanization procedure for sleeve deactivation that ensures a highly inert surface. These sleeves can be purchased from Restek at a cost below many instrument manufacturer's cost. Refer to Restek's Chromatography Products Catalog to obtain the part number for your specific instrument. For more information on inlet sleeves, request a copy of Restek's bulletin Operating Hints for Split/Splitless Injectors or Guide to Direct/On-column Flash Vaporization Injection. Protection Against Dirty Samples Sleeve packing materials such as fused silica wool (cat.# 20790), glass wool (cat.# 20789), or fused silica beads (cat.# 20791) act as filters when analyzing samples containing high molecular weight residue or particulates. However, both wool and beads greatly increase the surface area that the sample contacts and can be a source of adsorption or breakdown. It is critical that the wool or beads be properly deactivated. If you plan to use wool, be careful inserting it into the sleeve because active sites can be created as the fibers break. We do not recommend using packings coated with stationary phases. (For more information on protection against dirty samples, request a copy of Restek's bulletin A Guide When Injecting Dirty Samples.) Alternative sleeve designs that minimize sample interaction with non-volatile residue are also available. Replacing Critical Seals Replace the critical seal prior to installing an inlet sleeve (see instrument manual for seal location). Most capillary injection ports use a rubber o-ring or graphite ferrule to seal the sleeve inside the injection port body. The seal must fit tightly around the sleeve to prevent the carrier gas from leaking around the outside of the sleeve. If your GC uses a ferrule as the inlet seal, always pre-swage the ferrule to fit the sleeve before tightening it in the inlet (especially Varian inlets).  Changing Septa Always use a high quality, low bleed septa. We recommend replacing the septum frequently to prevent leaks and fragmentation. Otherwise, multiple injections and continuous exposure to a hot injection port will decompose the septum, causing particles to fall into the sleeve. Septum particles are a potential source of ghost peaks, loss of inertness, and carrier gas flow occlusion as shown in Figure F. It is best to install a new septum at the end of an analytical sequence so that it can condition in the injector and reduce the incidence of ghost peaks. Always use clean forceps when handling septa to avoid contamination. Setting Detector and Make-up Gas Flow Rates Confirm that the make-up gas, detector fuel, and oxidant flow rates are set according to the instrument's specifications (Table I, below). Make-up gas flow rates set too low will cause tailing solvent peaks, baseline disturbances, decreased sensitivity, and detector noise. Some instruments do not have leak-tight detector cavities and require flow rate verification before the column is installed into the detector. However, for GCs with leak-tight detector cavities, it is usually easier to check detector and make-up gas flow rates after the column is installed. We highly recommend using the Restek Flowmeter 6000 (cat.# 21622).
Table I Typical FID Flow Rates
| Instrument | H2 (ml/min.) | Make-up (ml/min.) | Air (ml/min.) | | Hewlett-Packard | 30 | 20 | 400 | | Varian | 30 | 20 | 300 | | Shimadzu | 30-60 | 40 | 500 | | PerkinElmer | 45 | -- | 450 | |
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Never use liquid leak detectors that contain soaps or surfactants in capillary chromatography. Leaks draw these materials inside the system and contaminate the column, making high sensitivity operation difficult. In addition, liquid leak detectors can cause permanent damage to the capillary column by depolymerizing the silicone stationary phase. 
Since compounds used to set the dead volume are usually gases at room temperature (methane), they are not extremely sensitive indicators of system or installation problems. A 1µl injection of a liquid solvent, such as methylene chloride, expands to over 500µl of gas volume, making any potential installation or system problem readily apparent. A tailing solvent peak is a sensitive indicator of broken, undeactivated, or contaminated inlet sleeves. Tailing solvents also indicate problems with inadequate make-up gas or improper column insertion into the detector. 
