ESPCI, PECSA UMR CNRS 7195, Paris, France. 

Supercritical fluid chromatography (SFC) is a valuable tool prior to comprehensive two-dimensional gas chromatography (GC×GC) analysis to investigate the composition of petroleum vacuum gas oils (VGOs) and selectively analyse phenolic compounds in coal-derived middle distillates. In the first case, a multidimensional SFC system is used to separate saturates, aromatics and polar compounds in a VGO sample, and in the second case a single ethylpyridine stationary phase is sufficient to selectively separate phenols.

Traditional fossil fuel sources are dwindling and new ways to isolate fuels from existing sources are vital. Two methods look particularly promising: converting heavy petroleum fractions into lighter fractions and transforming coal-derived liquids into synthetic fuels that fit engine requirements. These two matrices involve two separate problems. Converting heavy fractions, such as vacuum gas oils (VGOs), into specific groups is difficult because compounds contained in heavy petroleum fractions must be transformed into lighter compounds,

and coal-derived middle distillates have a high phenol content which must be precisely determined if these distillates are to be processed further (1). 

These problems can be solved by using a high-resolution technique, such as multidimensional chromatography, which was developed to overcome the limited peak capacity of one-dimensional chromatographic methods (2). 

Although comprehensive two-dimensional gas chromatography (GC×GC) (3) is popular in many sectors, including food, biology and environmental, it is still difficult to cope with some complex samples in terms of group-type selectivity and peak capacity. This difficulty has been highlighted in recent work on analysing diesel fuels (4) and heavy petroleum fractions, such as VGOs (5,6). The lack of group-type selectivity is often a result of the limited choice of polar stationary phases. To overcome this problem multidimensional systems incorporating a separation using a dense mobile phase have been suggested as a solution, including a SFC×GC system (7) and an LC×GC system (8–10).  

When considering on-line hyphenation of an LC system with a GC or GC×GC system, the key technical issue is the evaporation of the LC mobile phase which must induce a limited reinjection band for GC. 

The stop-flow mode via adapted valve-based- or syringe-based interfaces is preferred because of its flexibility (11,12). Although remarkable results have been obtained for edible oils (12) and petroleum samples (11), the total running time remains a major obstacle. 

 as the mobile phase seems more compatible with a gas phase dimension than an LC dimension. The improved compatibility of SFC can be attributed to the expansion stage from the supercritical state to the gas state without any loss of the products of interest. Additionally, SFC separations can provide higher selectivity because of the variety of solvent and solutes, and stationary phase interactions, and because they are often faster than LC separations (13).  

The hyphenation of SFC with GC has been already demonstrated by the decompression of the supercritical fluid via a restrictor inserted into the GC injection port (14). This was applied using SFC–GC (15). The symbol "–" defines a heart-cutting hyphenation between SFC and GC, and The "×" symbol in SFC×GC (16,17) defines a comprehensive coupling.  

Capillary SFC×GC was demonstrated by the pioneering work of Liu et al. (16). The effluent from the first SFC column was received continuously by the second GC column through a thermal desorption modulator (16). Venter et al. proposed a stop-flow SFC×GC system for oil samples analysis (7) with a fast GC dimension. 

An additional dimension involving SFC appears to be very complementary to GC×GC systems. 

A SFC–GC×GC (18) configuration was designed for the analysis of a diesel sample. The sample was separated into two fractions, which were directly transferred into two distinct GC×GC column sets (SFC–twin-GC×GC). However, the SFC fractions were not managed by a special interface. Therefore, an interface for multidimensional chromatography between SFC and GC was developed. The design provides an integrated solution for the hyphenation of a SFC system and GC×GC system, and can be operated as an SFC system, an SFC–GC system, an SFC–GC×GC system and also as an SFC×GC×GC system (19).  

This paper aims to show how this system was used to unravel the complexity of heavy VGOs and to separate phenolic compounds in coal-derived middle distillates. 

 The separation of vacuum gas oils by SFC was first studied with a synthetic test mixture prepared in carbon disulphide (Table 1). 

 Table 1: Composition of test mixture TM1. 

 A vacuum gas oil of an Arian light feedstock was supplied by IFP Energies Nouvelles (Lyon, France) (Table 2). Before injection, it was previously diluted in CS

Table 2: Physical characteristics and elementary composition of the VGO sample studied. 

Direct coal liquefaction middle distillate 

 To evaluate the potential of different columns to separate phenolic species, a model mixture representative of a coal- derived middle distillate was prepared. Six families of chemical were selected because they were identified in previous works (1,20): Paraffinic, naphthenic, aromatic, furanic, phenolic and nitrogen-containing compounds. For each chemical family a light and a heavy compound was selected (Table 3). The solvent used for the test samples was ethyl acetate and was supplied by VWR (Fontenay-sous-bois, France). Chemicals were provided by different suppliers: Sigma-Aldrich (Saint Quentin Fallavier, France), Merck (distributed by VWR), Alfa Aesar (Schiltigheim, France), TCI (Zwijndrecht, Belgium) and Fluka (Saint Quentin Fallavier, France).

Table 3: Model mixture representative of the coal-derived middle distillate. 

 The investigated coal-derived oil was provided by IFP Energies Nouvelles. It consisted of a middle distillate (boiling points range = 200–350 °C) of a coal oil produced by direct liquefaction. The sample used in this study had a hydrogen content of 11.4% 

 [by nuclear magnetic resonance (NMR) spectroscopy], a nitrogen content of 0.24% 

 (ASTM D4629), a sulphur content of 0.0072% 

 [provided by X-Ray Fluorescence (XRF)], and an oxygen content equal to 0.80% 

. The refractive index (RI) and density of the sample were 1.5129 (ASTM D1747) and 0.9330 g/cm

 Samples were injected in the instrument through a switching valve equipped with an injection loop of 5 µL. The mobile phase, carbon dioxide (Alphagaz CO

 SFC grade, Alphagaz, Mitry-Mory, France) was delivered by a PU-2080-CO

 pump (Jasco, Bouguenais, France) at a constant flow-rate of 2.5 mL/min. The backpressure was regulated by a BP-2080 pressure regulator (Jasco). The SFC column(s) and tubular connections were introduced in an in-house modified gas chromatograph (GC6890, Agilent Technologies, Massy, France). All two-positions switching valves were managed via a pneumatic controller (6 ports AC6W, VICI, Schenkon, Switzerland). All tubular connections were in stainless steel (1/16" × 0.13 mm, 0.005') (Interchrom, Montluçon, France). All of the valve heads were placed in regulated heat chambers located at the top of the GC chromatograph. The SFC separation was monitored with a flame ionization detector (FID) set at 350 °C. For this purpose, the total supercritical flow-rate was split (about 1:100) by an integral restrictor made in-house (21) using a stainless steel cross union ZT1C (1/16'', 0.25mm, VICI). The SFC was controlled by independent Chemstation software (Agilent Technologies).  

 The GC×GC system consisted of an in-house modified 6890 chromatograph (Agilent Technologies) equipped with dual-stage carbon dioxide jets (22). To perform  modulation simultaneously for two column sets (SFC–twin-GC×GC), the columns holder supplied by Thermo (Milan, Italy) was adapted on top of the chromatograph. The 2D modulation was performed at the beginning of the second columns and the modulation period was set at 30 s.  

The 2D modulation was managed by a controller built in-house. The GC×GC instrument was equipped with two split/splitless injectors and two FIDs (370 °C, 100 Hz). H

 and air flow-rates were 35 and 400 mL/min respectively. Two GC×GC columns sets were installed. Helium (99.99%) was used as the carrier gas at constant adapted pressure (23). During the transfer of SFC fractions into the GC×GC system, the trapping temperature was set at –30 °C.  

Once the SFC fractions were trapped on the GC phases, the oven returned to the initial temperature (100 °C for VGOs and 50 °C for coal-derived products) at 20 °C/min and the analytical ramp was controlled at 2 °C/min up to 370 °C for VGOs analysis and to 280 °C for coal- derived products analysis.  

The data processing of 2D contour plots was performed by 2DChrom (Thermo) after raw data transfer as CSV-files. The control of the (i) GC×GC dimension, (ii) the interface including the valves management and (iii) the data acquisition was performed using independent Chemstation software (Agilent Technologies).  

 An interface allowing the intermediate sampling of SFC fractions into loops was designed and implemented. The designed interface is shown in Figure 1. It should be noted that none, one, multiple or all of the supercritical effluents coming from the SFC part can be distributed into the GC×GC system. 

Figure 1: Schematic of the designed interface for  SFCâGCÃGCÃGC. 

The designed interface was introduced in a highcapacity oven supplied by Interchrom (Montluçon, France). To avoid any change in the physical state of the supercritical effluents, the same temperature and pressure conditions as the SFC separation ones were applied across the whole interface. Therefore, all of the tubular connections and the valve heads were placed in the oven using long metal actuators between the valve head in the oven and the motor out of the oven (VICI). 

This interface was based on a set of three main valves: V1, V2 and V3. The first two valves (6 ports AC6W, VICI) were managed using a pneumatic controller. V2 is a 12-port valve EMT2CST6UW (1/16", 0.4 mm, 0.016'), supplied by VICI and is resistant to high pressure (up to 345 bar) and temperature conditions (up to 75 °C); six HPLC sampling loops (model SLKCW, VICI) were adapted using the 12 ports of the multi-positions valve. The management of V2 was performed by a microelectric actuator (EMTCA-CE, VICI) directly piloted by the Chemstation software (Agilent Technologies).  

 The function of the V1 valve was the sampling management of supercritical effluents coming from the SFC part towards valve V3 (closed) or the multi-positions valve V2.  

V2 allowed the sampling or draining of supercritical effluents, by successively switching the rotor position, that is each loop, allowing for their transfer into or from a loop. The volume of the sampling loops depended on the SFC separation and the sampling of supercritical fractions. The optimal volume for the draining of loops was set at 1.5 times their physical volume and at a minimum of 10 mL.  

The V3 valve allowed the transfer of supercritical fractions into the GC×GC system, by splitting the total flow-rate using a derivation combined with a restrictor directly inserted in the GC×GC injector (14). V3 was closed when just implementing a single SFC separation, as well as performing a GC×GC run, that is fractions already transferred, to prevent leaks from the inlet septum. Supercritical fractions were expanded at the restrictor tip inside the GC injector, and then solutes were cryo-focused on the first centimetres of the first GC×GC column.  

After the transfer of each SFC fraction, the purge valve of the injector was opened to drain the residual CO

 out of the system. When two different columns sets were available in the same GC×GC oven for SFC–twin-GC×GC operation (two different injectors and detectors were used for GC×GC), for each fraction to be transferred from the SFC system, the selection of the GC×GC columns set to be connected to the SFC effluent was performed using another valve (V4) placed between V3 and the two restrictors.  

Concerning the transfer of supercritical effluents, the interface described by Levy et al. (14) has been implemented, that is the decompression of the supercritical fluid via a restrictor inserted into the GC injection port. Conditions were maintained at –30 °C for the trapping temperature, 350 °C for the GC injector temperature and 13 mL/min for the restrictor flow-rate. Supercritical mobile phase was depressurized using the integral restrictor before the trapping of solutes on the first centimetres of the cold first column. The splitless mode was used during transfer. Once the transfer was done, the purge valve was opened to release residual CO

 and the oven returned to the initial temperature (100 °C for heavy oils, 50 °C for coal-derived liquids) at a ramp of 20 °C/min; then the analytical ramp could start.  

 The selectivity of various stationary phases (Table 4) was studied with the test mixtures MT1 and MT2. Experiments were performed at 35 °C and the backpressure was regulated at 250 bar.

Table 4: Stationary phases studied for the SFC separation. 

 For the application for VGOs, a first 2D-GC column set was dedicated to SFC fractions composed of saturated compounds. It consisted of a combination of a non-polar first column: DB1-HT (dimethylpolysiloxane, J&W Scientific, Folsom, USA) (10 m × 0.32 mm, 0.1 µm) and a mid-polar second column, BPX-50, (50% phenyl)polysilphenylene-siloxane, (SGE, Courtaboeuf, France), (0.8 m × 0.1 mm × 0.1 µm). The second column set was specifically dedicated to SFC fractions composed of unsaturated and polar compounds. It consisted in a combination of a DB1-HT (10 m × 0.32 mm, 0.1 µm) and a BPX-50 (1.45 m × 0.1 mm × 0.1 µm). 

 For the analysis of phenolic compounds the column set consisted in a polar Solgelwax (30 m × 0.25 mm × 0.25 µm) in the first dimension coupled with a DB-1 (1 m × 0.1 mm × 0.1 µm) in the second dimension. This configuration involving a polar column in the first dimension and an apolar one in the second dimension was supposedly non-orthogonal because retention mechanisms of each dimension are not independant. However, this configuration enabled the separation of phenols from hydrocarbons, but these species could not be classified by carbon atom in the chromatogram in the same way as orthogonal configurations. 

 The implementation of a SFC separation before the GC×GC separation must improve on the lack of group-type selectivity observed for the high temperature 2D-GC (HT-2D-GC) analysis of VGOs (24). The resolution between saturated and unsaturated compounds and the selectivity by aromatic rings number needs to be improved in particular. Consequently, a stationary phase study was performed by one- and multidimensional SFC. 

 A "conventional" stationary phase [used in adsorption or normal phase (NP) chromatography] is selective enough to perform a group-type SFC separation of light matrices, such as gasoline (25) and even diesel oils (13,26) in saturates, monoaromatics or diaromatics. However, owing to a higher complexity, the chromatographic resolution can be reduced for some diesels (27). To check the possible applications of 1D-SFC to heavy fractions, such as VGOs, several promising selective stationary phases were studied. This was based on potential high π–π or dipolar interactions (28,29) by the stationary phases. After SFC–FID experiments on the test mixture TM1, chromatographic resolutions were calculated between different couples of compounds (Figure 2).

Articles

SFC–GCxGC is shown to be useful in fossil fuel analysis. Practical examples for the analysis of complex matrices are described.

SFC–GCxGC to Analyse Matrices from Petroleum and Coal

The 5th International Symposium on Recent Advances in Food Analysis (RAFA 2011)

Conference organizers, the Institute of Chemical Technology in Prague, the Institute of Food Safety (RIKILT) in The Netherlands and the International Association of Environmental Analytical Chemistry (IAEAC), would like to invite all food scientists from universities, research institutes, national and international agencies, governmental and commercial laboratories, industry and other food business-related companies to attend the 

5th International Symposium on Recent Advances in Food Analysis (RAFA) 2011 at the Clarion Congress Hotel Prague, Czech Republic, 1–4 November 2011.  

Novel analytical applications in food research and a routine analysis are required to meet quality control (QC) and food safety criteria, comply with legal and labelling requirements, determine nutritional value and to detect food fraud/adul teration. It should be noted that, in spite of the legislative efforts, food scandals occasionally occur and can originate from contamination from pesticide residues, veterinary drug residues, endocrine disruptors, processing contaminants, migration from packaging materials or natural sources (including mycotoxins and marine toxins). Additionally, since modern biotechnology methods have been introduced to food production, GM food and food ingredients have become an issue and have led to a strong demand for adequate methods of analysis.  

As a result the science of food analysis is a rapidly evolving and exciting field. In all cases, sampling strategy, sample analysis, data handling and final exposure assessment are crucial steps that are closely interrelated. The desired level of consumer protection and the strict regulation of maximum limits force analytical chemists in food laboratories to lower quantification limits, improve QA/QC and increase sample throughput by introducing new approaches and highly innovative technologies, including the use of bioassays for bioactivity and toxicity screening; omics-based analysis including metabolomics-like profiling; on-line and at-line analysis approaches; spectrometric food profiling (NMR, NIR); (bio)nanotechnology and chip-based methods; solvent-free and other emerging isolation technologies; and hyphenated techniques such as LC–MS and MS–MS, fast GC–TOF-MS, GC–MS–MS, ambient MS and/or comprehensive chromatography (GC?GC, LC?LC).  

At the last RAFA conference in 2009, 530 scientists from 55 countries participated. Fifty seven oral communications (including 13 young scientists') and 440 posters were presented within the scientific programme. Ten vendor seminars and two satellite workshops were also organised. Scientific committee and organizers of the 

 aim to prepare a balanced programme with high-quality presentations followed by stimulating discussions, several satellite events, a state-of-the-art exhibition and an attractive social programme.  

Scientific contributions will be presented by leading scientists through key note lectures and contributed oral and poster presentations. Moreover, a platform for young scientists to present their scientific work will be offered. 

 Student Travel Grants will be also provided. The best poster presentation(s) by a young scientist(s) will be awarded. Several satellite events are planned currently to accompany the conference programme. 

An exhibition of the modern instruments used in food analysis will be on display during the symposium. Vendor seminars will be also organized to introduce recent instrumentation and scientific strategies for advanced food quality/safety control. Oral presentations can be submitted until 

For further information contact Monika Tomaniova on +420 605 423 973, e-mail 

 Sherrington Building, University of Liverpool, UK 

5th International Conference on Packed Column SFC (SFC 2011)

The 17th International Symposium on Separation Sciences (17-ISSS)

Departments

Event News

In the second instalment of this series, John Hinshaw discusses the use of gas switching valves for column backflush and two-dimensional series-bypass applications.

Multidimensional gas chromatography (MDGC) as practiced today may be divided into two main application areas: Classical MDGC and comprehensive two-dimensional gas chromatography (GC×GC).  

Classical MDGC involves redirecting a portion, or portions, of the eluent from one column to another column with different separation characteristics, at times that are synchronized with the elution of specific peaks (or groups of peaks) from the first column.  

The more recent technique of GC×GC also involves the transfer of the eluent from one column to another, but the transfer occurs cyclically and continuously across the entire chromatogram without targeting the timing of individual peaks. All of the eluent from the first column is transferred to the second column so that all peaks are separated by both columns. The comprehensive approach yields resolving powers that approach the product of the first and second columns with a corresponding leap in data quantity and complexity. Increased knowledge and skills are required from the analyst to use this approach effectively in the routine laboratory.  

This month's "GC Connections" will describe some basic examples of classical MDGC and offers insights into the more advanced methods of GC×GC. 

The injection of gaseous samples from a fixed-volume loop or the swapping of detectors during a GC analysis are two simple examples of ways that chromatographers use gas switching valves (GSVs). 

The previous instalment (1) showed how these fundamental valve techniques work for packed and micropacked GC columns. Chromatographers also use valves in more complex ways, such as reversing the flow of carrier gas through a column, or to manipulate and redirect gas streams between two or more columns in multidimensional GC.  

MDGC techniques enable the separation of a wide range of components that might not be achievable with a single column or in a single analysis. Separating a wide range of compounds with conventional single-column GC may require column temperature programming across a relatively large temperature range while MDGC could provide an equivalent separation isothermally, sometimes in combination with carrier gas pressure programming.  

Dual-parallel-column analyses are possible, too, but sometimes it is not desirable to inject the entire sample onto both columns, especially if all injected substances cannot be cleared from both columns before a subsequent injection.  

Performing a pair of discrete separations one after the other is also possible, but this method would also place the entire sample into each column, as well as taking the sum of both analysis times to be completed.  

Before looking at multidimensional applications, we'll first take a look at a simple column flow-reversal application. 

Chromatographers use column backflushing when the sample contains residual materials that would not otherwise be eluted from a column until well after all peaks of interest have been eluted; as an alternative to high-temperature column oven excursions where the upper temperature limit may be curtailed by the maximum temperatures allowed for the valves or column; or if temperature cycling is not otherwise required. Column and valve lifetimes can be extended by avoiding repeated high temperatures over long time periods. Backflushing works by removing late-eluting impurities from a column that might otherwise interfere with subsequent analyses. 

Backflushing of a single column can be accomplished using the 6-port valve arrangement shown in Figure 1. In Figure 1(a) the 6-port valve starts in the backflush or standby position: carrier gas flow proceeds through the column from left to right as shown. The set-up is ready for injection after the sample fills the sample loop.

 Figure 1: Single-column backflush to detector configuration. (a) Standby/backflush position, (b) Injection/elution position. 

 When the valve is moved to position (b) the sample loop is flushed by carrier gas flow onto the right-hand column connection as the column flow direction reverses and starts going from right to left. As long as the sample loop pressure is low, there is no danger of the sample flowing backwards into the carrier gas pneumatics because the carrier gas pressure is higher than the low-pressure end of the column at valve port 2 at the moment of injection. 

The valve remains in the (b) position until the last peak of interest has been eluted. The valve is then returned to position (a), which causes the column flow to be restored to its original left-to-right direction (backwards in relation to the direction that the carrier gas flowed during peak elution). Any solutes or impurities left in the column at the backflush time will begin to move in reverse, towards the detector. As the total run time approaches twice the backflush time — assuming the carrier gas flow or pressure are not modified — residual material will be backflushed through the detector. At a time equal to approximately twice the backflush time nearly all of the material will have been flushed out. Some may remain as a result of band-broadening effects or nonlinear retention, so it is good practice not to perform another injection until sufficient additional time has passed in the backflush mode to remove all residual material. 

Figure 2(a) illustrates a backflushed chromatogram of a simple gas mixture. In Figure 2(b), the backflushed portion of the chromatogram is enlarged 200× to show a momentary baseline disturbance at the backflush time of 480 s, followed by some unidentified C

 and heavier compounds and a return to baseline before 960 s, at twice the backflushing time. Although the impurity levels are relatively low in this example, if they are not purged from the column after each run such extraneous non-eluted material could build up over many runs and cause baseline instability. Other examples include natural gas analysis with grouping of C

+ compounds, environmental extracts with high-boiling matrix compounds and food and beverage volatiles.

Figure 2: Column backflushing example. Conditions: 3 m Ã 1.0 mm i.d. porous polymer micropacked column; 70 Â°C isothermal; constant pressure; 1.0 mL injection; flame-ionization detection (FID). a) scaled to peaks; b) enlarged 200Ã. Backflush time: 480 s, indicated by arrow. Peaks: (1) methane, (2) ethylene, (3) acetylene, (4) ethane, (5) backflushed impurities. 

The example shown here was performed on a micropacked column. The principles of backflushing for open-tubular columns are similar, but because of the low dead-volume requirements such separations may be better managed with various fluidic capillary switching devices. A future instalment  of "GC Connections" will address these configurations and their applications.  

Some columns yield good separations for certain compounds but not for others. For example, a molecular sieves column discriminates between various light gases such as hydrogen, oxygen, nitrogen and carbon monoxide, but fails to separate water or carbon dioxide.  

These two compounds will contaminate a molecular sieves column until baked off at high temperatures, and if they are allowed to accumulate they will degrade the separation of other compounds on the molecular sieves column. On the other hand, certain porous polymer columns easily separate water and carbon dioxide as well as a variety of hydrocarbons and other gases, but may not produce acceptable separations of those gases handled well by molecular sieves.  

It is possible to combine the separation capabilities of these two columns for a complete separation of fixed gases and C

 hydrocarbons. The rest of this instalment will address the separation of these solutes in a couple of different ways. Other examples for different sets of solutes are easy to find in the chromatography literature. 

Two popular methods for this type of combined column two-dimensional separation are sequence-reversal and series-bypass operation. In both cases, the flow and connections between two columns are manipulated during the analysis via valve operations so that each column receives and separates the peaks that it is best suited for. 

The main differences between sequence reversal and series-bypass operations is that the former uses a single detector or two detectors in series, while the latter can use two discrete detectors for true dual simultaneous separations. Column sequence reversal is illustrated by the flow diagram in Figure 3; this configuration uses a 10-port valve. The details are not shown here for reasons of space, but several examples can be found on the manufacturers' websites. 

Figure 3: Flow diagram for column sequence reversal. (a) Injection/sequential flow and (b) Sequence reversal. See text for description of operation. Peaks: (1) carbon dioxide, (2) ethylene, (3) acetylene, (4) ethane, (5) impurities including water, (6) hydrogen, (7) oxygen, (8) nitrogen, (9) methane, (10) carbon monoxide. Note: the time scale for chromatograms increases from right to left. Chromatograms are shown with thermal conductivity detection to reveal the fixed gases. 

In sequence reversal some care must be taken to ensure that peaks from one column are not co-eluted with peaks from the other. Figure 3(a) illustrates the column flows in the first step of injection with sequential flow from column 1 to column 2.  

The sample is injected from the sample loop into column 1. The initial, partially-separated peaks from column 1 enter column 2 until those peaks destined to be separated on column 2 have been transferred. The valve is then moved to change to flow as shown in Figure 3(b); the sequence of the columns is reversed.  

Peaks remaining in column 1 continue to partition and move along the column until they are subsequently eluted and detected. Simultaneously, peaks from column 2 are eluted into column 1, pass through it with little to no additional retention, and enter the detector. Co-elution of peaks in a complex mixture such as in this example may be difficult to avoid. In some applications it may be necessary to add a second valve to stop flow in column 2 and delay its peaks from entering column 1 until the existing peaks in column 1 have been eluted. 

A related configuration keeps the second column flowing to the detector, but backflushes the first column to vent after the solutes to be separated on the second column have left the first column. An independent carrier gas source may be used for backflushing the first column. This configuration is sometimes called pre-column backflushing, and can also be found in manufacturers' literature. It is useful when only the peaks from the second column are to be analysed, and the remaining peaks in the first column can be discarded. 

For the application at hand, what we really want to accomplish is the simultaneous determination of all of the peaks. This can be made possible by combining the two applications discussed so far, backflushing and column-to-column flow redirection. The backflush application shown in Figure 1 is augmented by replacing the 6-port valve with a 10-port valve, and an additional 6-port valve, carrier gas source and detector are added. The 10-port valve provides the backflush functionality, and the 6-port valve controls the destination of the effluent from column 1 — either to column 2 or to detector 1 through a restrictor — while generating functionality similar to that of the sequence-reversal configuration of Figure 3. 

Figure 4 illustrates the flow paths of a series-bypass backflush configuration. In Figure 4(a) residual material from a previous injection is backflushed from column 1 to detector 1. The configuration remains in this position until the sample loop is filled and injection may be started. Figure 4(b) shows the flows for injection and series operation of column 1 into column 2. Here, the contents of the sample loop are injected into column 1 and the initial unresolved fixed-gas peaks plus methane are passed in series onto column 2. After the methane peak passes into column 2, the valves are repositioned to produce the flow paths shown in Figure 4(c). 

Figure 4: Flow diagram for series-bypass operation with pre-column backflush. Discrete detectors for each column channel eliminate peak overlaps. (a) Standby/backflush flow. (b) Inject/series flow, (c) Bypass flow, Peaks as labelled in Figure 3. Note: the time scale for chromatograms increases from right to left. Chromatograms are shown with thermal conductivity detection to reveal the fixed gases. 

Column 1 now bypasses column 2, and both columns flow to their respective detectors. The solutes that were initially passed from column 1 to column 2 are eluted into detector 2, while the remaining target peaks from column 1 are eluted into detector 1. After the column 1 target peaks have been eluted, the flows are reset to position (a), so that any remaining contaminants in column 1 are backflushed to detector 1. The restrictor, which has similar flow characteristics to column 2, serves to maintain a more constant flow in column 1 as it is switched in front of column 2 and back again.

Figure 5: Series-bypass with backflush valve diagram. Both valves shown in the A position, for standby/backflush operation. See Table 1 for the relationship of valve positions to the flow paths shown in Figure 4. 

The valve plumbing for the series-bypass backflush configuration is shown in Figure 5. The 10-port valve provides sample injection and backflush functionality, while the 6-port valve selects whether column 1 effluent is directed to column 2 or to detector 1 via the restrictor. The valve positions in Figure 5 that correlate with the three flow configurations of Figure 4 are listed in Table 1.

Table 1: Valve positions for Figure 5 to achieve flow paths diagrammed in Figure 4. 

The timing of the bypass operation is important. If the first column is bypassed around the second column too soon, then some of the peaks that should enter the second column do not. If the bypass time is set too long, then some of the peaks that should remain in the first column can enter the second column instead. Figure 6 shows the effects of increasing the bypass time by 20 s. The methane peak is split between the first and second columns with a 60-s bypass time. Increasing the bypass time by 20 s results in all of the methane appearing on the second column. This timing must be revalidated any time the flow, temperatures or the columns themselves are changed. The timing of the backflush operation, at 480 s in this example, is not as critical and can occur at any time that the transient spike does not interfere with a peak, once all of the peaks of interest have been eluted from column 1.

Figure 6: Tuning the bypass time. (a) Porous polymer column with flame-ionization detection (column 1 in Figures 4 and 5), (b) Molecular sieves 5A column with thermal-conductivity detection (column 2 in Figures 4 and 5). The methane peak (9) is split between the two columns at a bypass time of 60 seconds, and is passed entirely to the molecular sieves column at a bypass time of 80 s. Peak labels as in Figure 3. 

This month's instalment examined the basics of valve applications for backflushing, sequence reversal and pre-column bypassing. We then combined these basic techniques to create a hybrid dual-valve configuration capable of separating both fixed gases and hydrocarbons in a single two-dimensional analysis. MDGC presents a powerful suite of techniques that analytical chemists can use to tease apart solutes that might otherwise be inseparable, at least without significant additional effort. The next instalment in this series will discuss multidimensional separations as applied to open-tubular columns; including fluidic flow switching and applications to heart-cutting, selectivity tuning of serially connected columns and GC×GC separations. 

 is senior research scientist at BPL Global Ltd, Hillsboro, Oregon, USA and a member of 

 editorial advisory board. Direct correspondence about this column should go to 

 editor, Alasdair Matheson, 4A Bridgegate Pavilion, Chester Business Park, Wrexham Road, Chester, CH4 9QH UK, or e-mail: 

GC Connections

The 2nd part of this series discusses the use of gas switching valves for column backflushing and 2D series-bypass applications.

Valves for Gas Chromatography (Part 2): Applications

Where do you start when you encounter a problem with your LC system? 

Old timers, like me, find that troubleshooting a liquid chromatography (LC) problem is instinctive. But this is not in our DNA — we had to learn these skills over time. I continually see new faces in the LC world who are not as fortunate as I was. They do not have mentors to show them the ropes. So I would like to go back to the basics of LC troubleshooting and consider some fundamental problem-solving approaches that we can apply to most LC problems. In this month's "LC Troubleshooting" we'll start with some general principles. In future instalments, we'll look at specific problems in more detail.  

If you consider all the LC problems that you are likely to encounter, they generally fall into one (or more) of five categories:  

Peaks that are out of place in the chromatogram

One of my six troubleshooting rules of thumb is called "The Divide and Conquer Rule". The way I apply this is to make tests or observations that eliminate potential problem sources. That means I start by classifying the problem into one of the five categories listed above, so that we don't have to worry about the other categories. Let's look at each of the categories briefly. 

You will notice that any LC method that you use has a normal system pressure or system-pressure range. For example, an isocratic method might normally run at approximately 150 bar (approximately 2200 psi); or a gradient method might have a pressure range during the gradient of 140–200 bar (2000–2900 psi). When the pressure increases or decreases 20% from the normal values, then perhaps the system is trying to tell you something. You may notice the pressure change when examining the system, or the high- or low-pressurelimit setting may trigger an alarm or a system shut-down. Pressure can rise, fall or fluctuate. Each of these factors can tell you something about potential problems, such as the presence of blocked frits, leaks, bubbles, or worn parts.  

Leaks are often one of the simplest problems to isolate, but not always. Many LC systems have leak detectors consisting of simple electrical contacts that are connected by a few drops of conductive mobile phase when a leak occurs. In other cases, you may notice a puddle of mobile phase on the bench or a buildup of white crystals at a fitting. 

Under normal operating conditions, the peaks in a chromatogram should have constant retention times. There will often be a little variation from run to run (generally 0.1 min or less, or perhaps a little longer for day-to-day variations), but in each case, a normal pattern of variation will occur. When the pattern is broken, it is a sign that something is wrong. Sometimes you will observe a drift for some (or all) of the peaks to longer or shorter retention times. In other cases, peaks may be missing, or extra peaks may appear. These can be attributed to problems with the sample, the column, the mobile phase or the LC hardware. This category of problems is perhaps the most common one and the most difficult to troubleshoot. 

Ideally, peaks in chromatograms will be symmetrical, but most peaks have a little bit of a tail on them. When this tailing increases or when peak fronting appears, something is going wrong. Modern columns, which use highpurity silica packings, are much less susceptible to peak tailing problems than the lowpurity columns that were standard 15–20 years ago. But you may be using a method that was developed on an older column, so tailing may be an everyday occurrence for your method. A change in peak tailing is often associated with a problem with the mobile phase or column. 

Sometimes you will first notice a problem when you are examining the data produced from a batch of samples. Perhaps the quality-control (QC) samples are out of range; the calibration curve is no longer linear; or all the peaks are too large or too small. Problems of this nature can often be traced to sample preparation or stability; or to problems with the autosampler. 

Sometimes you will find a problem that fits into more than one category. For example, a leak will cause the pressure to drop; the flow-rate will then also decrease, which will increase the retention times of the peaks in the chromatogram. Conversely, a leak may occur because of a blockage, in which case the pressure will go up. Because some problems fall into more than one of the five categories above, it is a good idea to determine if this is the case, because it may simplify the troubleshooting process. For example, if you notice increased retention times, look for leaks. This is a quick check and leaks are usually easy to fix. If you ignore the possibility of a leak and focus on mobile-phase problems, you might take much longer to find the root cause of the problem. As we look at the potential causes of the various symptoms, I'll try to show you how you can consider any one of the multiple categories for a problem and quickly find the source of the problem. 

I have six rules of thumb to help me remember some key troubleshooting techniques:  

Let's look at each of these in a little more detail: 

 "The Rule of One" says to change just one thing at a time. This is the scientific method, which we use in most of our work in the laboratory, but we seem to forget it when we are troubleshooting an LC problem. For example, if peak shape is poor, you might change the column, the guard column and the mobile phase to fix the problem. But if you change all three of these at once, you don't gather information that will help you to identify the problem the next time it occurs and shorten the next troubleshooting cycle. So change one thing, then another, until you find the root cause of the problem.  

Of course, there are times when it may be appropriate to ignore the "The Rule of One," but these should be the exception, rather than the rule. For example, if it is 4 pm on Friday and you have the problem mentioned above, it may be faster to change several things to help ensure that you can get the system operating for a long run over the weekend. 

 "The Rule of Two" tells us to make sure that we have a problem that is reproducible, that is, it occurs at least two times. Unfortunately, some problems with an LC system or method may happen once in a while, but not regularly enough to easily isolate and correct. For example, when examining the data after a batch of samples, we notice that a quality control (QC) sample has an area that is unexpectedly low. Is this a real problem that should be fixed, or did something happen to that one injection that is a random error? The easiest way to apply "The Rule of Two" is to repeat an injection (or inspect duplicate injections that are part of an existing data set). Do you get the same result twice? If the result is reproducible, you now have a change that you can focus on. Does the same low peak area occur for all QC samples or just some? How about other types of samples such as test samples or calibration samples? You can use "The Rule of Two" to help you begin to find failure patterns, and once a pattern is observed, tracing its origin is much easier. 

 "The Divide and Conquer Rule" rule was briefly mentioned earlier. I try to make an observation or experiment that allows me to discard a large number of possible root causes of a problem. Then I divide the territory again and again until I find the problem source. Does the problem occur only with samples, but not with standards? Does the problem occur more often with a column that has had 1000 samples through it than with a fairly new column? Does it show up at the beginning of a sample batch, but disappear by the end? Is it specific to one model of LC system, one operator or one method? By eliminating the things we know are 

 the cause of the problem, we can more easily identify those that 

 "The Module Substitution Rule" is perhaps the most powerful way to isolate the source of a problem. This simply involves replacing a suspect part with a good one. The most common application of module substitution is exchanging one column for another, or replacing a batch of mobile phase with a fresh one. Module substitution can be practiced at any level, from exchanging an entire LC system to a circuit board or component on a circuit board. In the context of hardware troubleshooting, the power of module substitution may help justify certain instrument purchase decisions. For example, if you are budgeted to purchase a new LC system, seriously consider getting another system just like the one you have. Then you can exchange parts — pumps, autosamplers, detectors and so forth — to isolate problems much more easily than if the two systems were different brands. You will also be able to transfer specific troubleshooting and maintenance skills more readily. 

 Sometimes I include "The Put-It-Back Rule" in the list of rules of thumb. This is closely related to "The Module Substitution Rule". It reminds us to put a good part back into service if it was replaced during module substitution, but was not found to be faulty. Although this process may seem obvious, it is amazing how many columns are discarded in this manner. You may replace the column as part of the problem-isolation process, but later realize that the problem was related to the mobile-phase pH, not the column. But you are in a hurry to get going, so you start the next sample batch with the new column. The original column, although perfectly good, gets set aside, and is often forgotten. How likely are you to pick up that column and use it as a replacement for another column in the future? Usually a used part is suspect and we'd rather not risk using it, so we discarded it. A good column-tracking procedure, where the sample history for each column is recorded, might help to avoid this problem. But as a general practice, if a new part substituted for an old one doesn't fix the problem, reinstall the old part. 

 One of my friends who works in quality assurance (QA) has a favourite saying: "If it isn't documented, it didn't happen." Most of the readers of this column work in the pharmaceutical industry, where good record keeping is mandated by the regulatory agencies. But besides being a good defence to use during an audit, troubleshooting and maintenance records can be invaluable tools to help avoid future problems. One of the uses of these records is to establish failure patterns or lifetime expectations for various components of an LC system or method. For example, if we wait until the pump seals fail, there can be a number of secondary problems, such as having to reanalyse a batch of samples that did not pass because of flow variations. In addition, the frit at the top of the column may get blocked from the bits of seal material that are sloughed off during the wear process. 

For those of you using ultrahigh pressure liquid chromatography (UHPLC) systems, you might consider column frit blockage a lucky occurrence: the extremely small-diameter tubing in the system may get blocked before the pump seal fragments reach the column. The expense of a new column and the hassle of flushing or replacing tubing might be avoided if you had a policy for pump seal replacement. For example, after several failure cycles, you may find that seals tend to fail between six and twelve months of use. Why not replace the seals proactively every six months and avoid the problem completely? Although pump seals are not inexpensive, they are less than 10% of the cost of a column — and all it takes is one failed seal to ruin an otherwise good column. 

Whether you decide to keep your LC maintenance and troubleshooting records electronically or on paper, be sure the process is simple and easy to use. Otherwise, good intentions may get ignored because of the hassle factor. My favourite recordkeeping system is still a looseleaf notebook, one for each LC system. With separate sections in the notebook for maintenance, troubleshooting, performance tests and so forth, the data are easy to locate when they are needed. A simple form may prompt for the 5 W's and the H of a good news story: who, what, when, where, why and how. Just a few words to summarize are all that are needed. Be sure to record part numbers and serial numbers, where appropriate. Armed with this kind of record for each system, it is easy to sort through different troubleshooting episodes and find common causes that may lead to preventive maintenance strategies. 

We have looked at the general problem categories that encompass most LC problems that you will encounter. We have also considered several rules of thumb that can be applied to many different types of problems to help isolate a problem, determine its source and, perhaps, develop a preventive maintenance routine that will help minimize the possibility of future downtime. In future instalments of "LC Troubleshooting" we'll look at some of the problem categories in more detail. 

 is vice president of LC Resources, Walnut Creek, California, USA. He is also a member of 

, editor, Alasdair Matheson, 4A Bridgegate Pavilion, Chester Business Park, Wrexham Road, Chester CH4 9QH, UK, or e-mail 

Where do you start when you encounter a problem with your LC system?

Back to Basics (Part 1): Where to Start?

The benefits of using software to optimize GC methods with practical examples for pesticide analysis are described in this month's "Column Watch".

Everyone wants to improve productivity in their chromatography laboratory. In this month's "Column Watch", we will show how a software approach can be used in gas chromatography (GC) to shorten analysis time but preserve the elution order and the resolution. The calculations involved are a bit more complex than for LC because the mobile phase in GC is a gas, which is compressible, while LC solvents are less so. In both techniques the mobile phase viscosity is temperature-dependent. 

There are various ways to reduce analysis time in GC:  					

Increase temperature programme ramp rates

Use hydrogen as a carrier gas (rather than helium or nitrogen)

Like any chromatographic method, there are trade-offs in any attempt to decrease analysis time. A balance between speed, capacity and resolution must be selected for each analysis to meet the laboratory's goals. Adapting methods for fast GC analysis can be complicated because peak reversals can occur and some fast GC methods may even decrease separation efficiency. Table 1 compares the benefits and drawbacks when changing to faster GC methods. 

Table 1: Benefits and drawbacks when changing to fast GC methods. Adapted from Reference 1.			

The GC Method Translation Software (1,2) is similar to the LC Method Translator, which will be discussed in the next column. This programme allows you to transfer a current GC method to another while ensuring that the relative retention order is maintained, that is, the peaks elute in the same order. It is free, stand-alone software that runs on a PC and will properly scale factors such as gas velocity and the temperature programme to make sure that the elution order is preserved. Users can easily translate to different column dimensions, faster flow-rates, switch carrier gases or a combination of all of these. One can do something as simple as change from a flame ionization detector (FID) to a mass spectrometry (MS) detector (operating under vacuum) or something as complex as changing the column length, i.d., detector type, carrier gas type, phase ratio (film thickness) and flow-rate (head pressure). The GC Method Translation software will generate a new method that will recommend a temperature programme, head pressure, and other parameter settings. The elution order will remain the same and the new method will attempt to maintain the resolution and selectivity of the original method. Peaks in the translated method do not have to be painstakingly identified. Thus, the GC Translation Software will reduce method development time or time to validate the new method. Furthermore, it will ensure that the new GC method is compatible with the hardware being used. Experimental parameters can be easily "tweaked" to speed-up the run time.  

A couple of caveats should be considered when using the GC Method Translation Software. First, you must not change the stationary phase because the software cannot correct for changes in selectivity. Columns with 100% methylpolysiloxane or 5% phenylmethylpolysiloxane from different manufacturers can often be used interchangeably but more polar phases can vary significantly between manufacturers. Note that as a column "ages", the stationary phase may suffer oxygen damage at elevated temperatures or become contaminated with non-volatiles that affect peak elution order initially and over time. Also, if you are using flow programming, this parameter is not addressed in the software. You could investigate various flow-rates using the Method Translation Software and see how changes in flow may affect other parameters. After collecting data at several different flow-rates, the best set of conditions can be selected for the application. 

The Method Translation tool operates in several modes: 

 Translates the current method to a new one based on a change of column dimensions, carrier gas type, outlet pressure and/or phase ratio. The elution order is preserved and the relative efficiency of the current method is maintained. This is useful for converting an established method to a column with different dimensions or a different phase ratio, or to change a detector with a different outlet pressure (e.g., a mass spectrometer). 

 Calculates new conditions (using your current column) that correspond to the theoretical optimum gas flow-rate for the best separation efficiency. The elution order remains the same but retention times may change. 

 Calculates temperature and pressure for your current column and carrier gas for a run that is twice as fast as the "Best Efficiency" mode; depending on the pressure drop across the column, run time will decrease by a factor of 1.5–2. The elution order of compounds will stay the same. 

 Allows you to change any parameter of interest; you can play with column dimensions, flowrate, head pressure, temperature programme rate and other parameters — even before you actually do any chromatography. You can use it with your current method to see how some parameters affect your results. For example, you can enter a smaller i.d. column to see the impact on run time; if your current method already has plenty of resolution, you can calculate optimum conditions for a shorter column operated at a higher flowrate and a different carrier gas. Thus, you can get a feel for the effect of each parameter adjustment on your chromatography. 

The GC Method Translation Software is easy to implement. It allows one to make simple or complex changes in a GC method to speed-up the analysis, change column dimensions, film thickness, use different detectors, choose a different carrier gas and a multitude of other changes. Without running an experiment, one can actually make hypothetical changes and see how they affect the chromatography and help to determine if the proposed changes can be implemented with the current instrumentation. Although GC Method Translation Software can be used for method development, it is better served when one wants to improve or change an existing method. A major factor in using this tool is that the elution order remains the same, so one does not have to go back and identify each peak in the chromatogram. 

Figure 1: Capillary GC analysis of solvents in pharmaceuticals. Original chromatographic conditions (a): Column: DB-624, 30 m Ã 0.25 mm Ã 1.4 Âµm; carrier gas: Helium; injection temperature: 225 Â°C; pressure: 16.2 psi; split ratio: 25; split flow: 37; total flow: 42.2; flow: 1.5 mL/min; mode: constant flow; Oven temperature programme: 40 Â°C for 3.84 min, then 40â200 Â°C at 13.01Â°C/min, hold at 200 Â°C for 1.87 min; detector: FID, hydrogen flow: 40 mL/min; air flow: 400 mL/min; make-up flow: 20 mL/min. Translated chromatographic conditions, (b): Carrier gas: hydrogen; see Figure 2 for calculated method parameters. 			

We will illustrate how this works in practice with two examples: one very simple, one a bit more complex. Both screen captures and actual chromatograms will be used to show how to use this tool. The first example involves a complex sample mixture and simply changing the carrier gas. The sample mixture consists of potential residual solvents in pharmaceutical formulations, typical of the United States Pharmacopeia (USP) method 467.(4) Our initial chromatogram and chromatographic conditions are depicted in Figure 1(a). A screen capture of the GC Method Translation Software is shown in Figure 2. The chromatographic parameters under consideration are shown in the left hand column of the screen while the initial conditions of the original method are shown in the second column. Here we used a DB-624 column (Agilent Technologies, Santa Clara, California, USA) with a stationary phase of 6% cyanophenylpropyl/94% dimethylpolysiloxane. The carrier gas was helium and an FID was used.  

Figure 2: Screen capture: Solvents in pharmaceuticals. 			

Using a temperature-programmed run, a baseline separation was achieved in approximately 15 min on the 30 m × 0.25 mm i.d. column with a 1.4 µm film thickness. It was desirable to speed-up the separation without having to change the column or affect this nice resolution. Switching to hydrogen carrier gas, the right hand column of the screen capture in Figure 2 shows the conditions recommended by the Method Translation Software. Making these recommended changes in flow-rate and in the temperature programme, the chromatogram shown in Figure 1(b) now provides a 10 min baseline separation of the 30 solvents with little or no change in resolution. The entire new method was set up by the Method Translation Software without the user performing any manual calculations and without a change in the elution order of the solvent peaks. Other changes could be investigated without running another chromatogram such as increasing the flow-rate or shortening the column. 

The second example involves systematic method changes looking at several chromatographic parameters. The sample this time is a mixture of 20 organochloro pesticides (OCPs). The determination of OCPs in environmental remediation samples is important, involving high-volume analyses in a competitive contract laboratory marketplace. A standard Contract Laboratory Protocol (CLP) pesticide method was used for these analyses. In many cases a laboratory will analyse large numbers of samples over the course of a given project, adding costs to both the laboratory and its client. It was desirable to decrease the analysis time by changing the carrier gas from helium to hydrogen and also decreasing the column diameter and shortening the column (but keeping the phase ratio the same). 

Figure 3: CLP pesticide method translation results. (a) Column: DB-35ms UI, 30 m Ã 0.25 mm i.d. 0.25 Âµm; carrier: He, constant flow, 45.17 cm/s at 110 Â°C; injector: splitless, 250 Â°C, ultra inert single taper liner with wool; oven: 110 Â°C for 0.638 min, 11.8 Â°C/min to 320 Â°C, hold time: 2.55 min; detector: Âµ-ECD, 330 Â°C, N2 makeup gas at 60 mL/min, (b) Column: DB-35ms UI, 20 m Ã 0.18 mm i.d. 0.18Âµm, carrier: He, constant flow, 50.0 cm/s at 110 Â°C; Injector: splitless 250 Â°C, ultra inert single taper liner with wool; oven: 110 Â°C for 0.384 min; 19.54 Â°C/min to 320 Â°C, hold time: 3.0 min; detector: Âµ-ECD, 320 Â°C, N2 makeup gas at 60 mL/min. (c) Column: DB-35ms UI, 20 m Ã 0.18 mm i.d. 0.18 Âµm; carrier: H2, constant flow, 78.2 cm/s at 110 Â°C; injector: splitless 250 Â°C, ultra inert single taper liner with wool; oven: 110 Â°C for 0.246 min, 30.55 Â°C/min to 320 Â°C, hold time: 2.0 min; detector: Âµ-ECD, 320 Â°C, N2 makeup gas at 60 mL/min. 			

Figure 3(a) shows the original method accomplished using DB-35ms UI (Ultra Inert) (Agilent Technologies), a 35% phenylmethylpolysiloxane phase designed to determine trace polar compounds, such as pesticides. High-content phenyl phases are known to have high selectivity for OCPs. As a result of its arylene backbone, the phase is also known for its thermal stability and high upper temperature limit (360 °C). The baseline separation of the 20 OCPs was accomplished in just over 18 min using a helium carrier gas and temperature programming conditions. A highly sensitive and halogen-selective electron capture detector (uECD) was used. The column dimensions were 30 m × 0.25 mm i.d. with a 0.25 µm film thickness. The first change investigated was moving to a shorter, smaller internal i.d. column with the same stationary phase. In GC, column efficiency is enhanced by using smaller i.d. columns at the expense of pressure increase. It is necessary to be aware of the column pressure at the high point of the temperature programme to avoid exceeding the rated pressure for the GC inlet. 

Figure 4: Screen capture: CLP pesticides method translation from 250 Âµm i.d. to 180 Âµm i.d. column using He carrier gas. 			

Typical gas chromatographs are usually pressure rated up to 100 psi. If this pressure is not exceeded, a method can be successfully translated to a smaller i.d. column. In our case, we also wanted to speed up the separation and a new, shorter column dimension was chosen (20 m). To keep the phase ratio the same the film thickness of the new column was estimated to be 0.18 µm. The GC Method Translation Software determines the optimum film thickness for commonly available in-stock columns for most manufacturers. Using the Translate Only mode of the software, Figure 4 shows the suggested experimental parameters for a new method, including column head pressure. The chromatogram in Figure 3(b) shows that using the suggested conditions and column (keeping He as the carrier gas), the separation time was reduced to just under 11 min: a saving of 7 min compared with the original method. 

Figure 5: Screen capture: CLP pesticides method translation from 250 Âµm i.d. to 180 Âµm i.d. column using hydrogen carrier gas. 			

The second parameter investigated was a change in carrier gas from He to H

. As was shown in the first example, a further decrease in analysis time was determined by the GC Method Translation Software (see Figure 5 for the screen capture showing the original method and the new translated method involving both column dimensional changes and carrier gas changes). The chromatogram for the new method is depicted in Figure 3(c). Thus, a further reduction of the separation time to approximately 7 min gave a 60% reduction in time from the original method. Figure 6 shows a direct comparison of the chromatograms, emphasizing the equivalent resolution of the 22 CLP pesticide peaks under all three sets of conditions. 

Figure 6: Comparison the of same three chromatographic conditions as Figure 4: Faster elution, same resolution. 			

The GC Method Translation Software is a free package available online or to download that helps the user to make changes to a GC method. Two examples were provided in this instalment of "Column Watch" to show how easy it is to use this software to convert, speed up and improve GC methods by changing the carrier gas and column dimensions. The tool can be used to simulate changes to other chromatographic conditions — even without performing any experiments. It also suggests experimental parameters within the scope of the instrumentation being used. For example, if you want to speed up a separation using a 0.05 mm i.d. column at a high flow-rate, the method translator would determine if the pressure required would exceed the 100 psi available from the standard gas chromatograph. 

 is a senior applications chemist in the Chemistries and Supplies Division of the Chemical Analysis Group at Agilent Little Falls Site, Wilmington, Delaware, USA. He has worked in the pharmaceutical, environmental and industrial chemical sectors for more than 20 years developing applications using GC, GC–MS, HPLC and SFC techniques. His current focus is on application support for Agilent's capillary GC columns.  "Column Watch" editor, 

, is a senior scientist at the Columns and Supplies Division. Agilent Technologies, Wilmington, Delaware, USA and is a member of 

 editorial advisory board. Direct correspondence about this column should be addressed to "Column Watch", 

, 4A Bridgegate Pavilion, Chester Business Park, Wrexham Road, Chester CH4 9QH, UK or e-mail the editor, Alasdair Matheson, at 

2. M.S. Klee and V. Giarrocco, "Predictable Translation of Capillary Methods for Fast GC", Agilent Technologies Application Note 5965-7673E, March, 2000. 

http://www.chem.agilent.com/en-US/Support/Downloads/Utilities/Pages/GcMethodTranslation.aspx

4. USP general chapter <467>, Residual Solvents USP 30/NF 25 procedure, 2nd supplement, effective July 2008. 

Column Watch

The advantages of using software to optimize GC methods with practical examples for pesticide analysis are described.

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