The Two Options for Sample Evaporation in Hot GC Injectors

Partial solvent evaporation inside the syringe needle (optimized as “hot needle injection”) produces thermospray: the sample liquid ... Lingfei Ho...
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Anal. Chem. 2002, 74, 10-16

The Two Options for Sample Evaporation in Hot GC Injectors: Thermospray and Band Formation. Optimization of Conditions and Injector Design Koni Grob* and Maurus Biedermann

Official Food Control Authority of the Canton of Zurich, P.O. Box, CH-8030,Zurich, Switzerland

Although classical split and splitless injection is more than 30 years old, we only start to understand the vaporization process in the injector. Solvent evaporation determines much of the process and is the first obstacle to overcome. Videos recorded on devices imitating injectors showed that sample (solvent) evaporation is often a violent process which is poorly controlled and might well explain many of the puzzling quantitative results often obtained. We do not adequately take into account that two vaporization techniques are in use. Partial solvent evaporation inside the syringe needle (optimized as “hot needle injection”) produces thermospray: the sample liquid is nebulized upon leaving the needle. The resulting fog is rapidly slowed and moves with the gas. Solute evaporation largely occurs from microparticles suspended in the gas phase. Empty liners are most suitable. Fast autosamplers suppress vaporization in the needle, i.e., nebulization, and shoot a band of liquid into the chamber that must be stopped by a packing or obstacles suitable to hold the liquid in place during the 0.2-5 s required for solvent evaporation. Solute evaporation largely occurs from the surfaces onto which the sample is deposited. Insights into these mechanisms help optimize conditions in a more rational manner. Methods should specify whether they were optimized and validated for injection with thermospray or band formation. The insights should also enable a significant improvement of the injector design, particularly for splitless injection. Split and splitless injection are the most widely used techniques of sample introduction in capillary GC, despite numerous inherent problems and rather frequent disappointing quantitative performance.1 The sample, usually a liquid, is introduced into a hot chamber in order to convert it to a vapor. The gas flow carries the vapor to the column entrance positioned at the bottom of the chamber. When introduced in split mode, a usually small proportion is driven into the column, whereas the rest is discharged through the split outlet. In splitless injection, the split exit is closed during a time that is long enough to transfer almost all sample vapor into the column and then reopened to flush the injector. * Corresponding author:-(e-mail) [email protected]; (fax) +41 1 262 47 53. (1) Grob, K. Split and Splitless Injection in Capillary GC; Wiley/VCH: Weinheim, 2001.

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Classical split and splitless injection still struggle with basic problems, such as poor control of the vaporization process (e.g., uncertainty on when and how to pack the liner), overfilling of the injector chamber, uncontrolled expansion of the vapor resulting from the pressure increase during solvent evaporation, and insufficient understanding of matrix effects.2 There seems to be a lack of coherence, i.e., of convincing concepts and rules on optimizing conditions. These problems may result in high standard deviations, systematic errors, and puzzling quantitative performance and are the main reason for the reputation of capillary GC to be excellent in separation but poor in quantitation. Recent progress in understanding of the vaporization process should enable a major step forward and help to get the techniques out of the deadlock in which they have been trapped for more than a decade. SAMPLE EVAPORATION Solvent Evaporation as First Obstacle. A rough estimation of the amount of energy consumed by sample evaporation is sufficient to conclude that vaporization cannot be the instant and smooth process we tend to assume.3 The processes taking place in classical split/splitless injectors were videotaped, using devices imitating vaporizing chambers and perylene, a fluorescent compound as long as in liquid phase (solution), as a marker for nonevaporated sample.4 This provided a wealth of new insights.5,6 Many observations were alarmingsin the end it seemed rather surprising that quantitative results are, nevertheless, as reproducible and accurate as commonly observed. Particularly the role of the solvent in the evaporation process was probably underestimated. (1) Solvent evaporation is the first obstacle to overcome since solute evaporation only starts afterward. Droplets of sample remain at a temperature near the solvent boiling point until solvent is fully vaporized. (2) Solvent evaporation is time-consuming (around 0.2-5 s) because of the large heat consumption involved. The liquid must be held in an appropriate position during this time (see below). (2) Grob, K. Anal. Chem. 1994, 66, 1009A-10019A. (3) Grob, K. J. High Resolut. Chromatogr. 1992, 15, 190-194. (4) Biedermann, M. Visualization of the Evaporation Process during Classical Split and Splitless Injection in GC, CD-ROM; Restek Corp.: Bellefonte, PA, 2000. (5) Grob, K.; Biedermann, M. J. Chromatogr., A 2000, 897, 237-246. (6) Grob, K.; Biedermann, M. J. Chromatogr., A 2000, 897, 247-258. 10.1021/ac0107554 CCC: $22.00

© 2002 American Chemical Society Published on Web 12/01/2001

(3) Partial solvent evaporation in the syringe needle nebulizes the sample liquid at the needle exit and provides a totally different starting point for solute evaporation than fast autosampler injection shooting a band of liquid through the chamber. (4) Solvent evaporation in the injector is responsible for sometimes violent movements of the liquid and exploding droplets and thus determines the distribution of the solute material in the vaporizing chamber. (5) The solvent determines whether the solute material is deposited on to surfaces, which largely determines matrix effects. (6) Usually over 99.9% of the vapor is from the solvent. The large volume of solvent vapor causes the problems regarding injector overloadingsor, seen from another angle, compels us to use large vaporizing chambers. In conclusion, often solvent evaporation should be improved when solute evaporation causes problems. The Two Principal Ways of Sample Evaporation. For understanding of the solvent evaporation process in the injector, it is essential to take into account the behavior of the sample liquid in the syringe needle when the latter is inserted into the hot injector chamber. With classical (in particular, manual) injection, volatile solvents start evaporating inside the needle before the plunger of the syringe is depressed. This was recognized as an important source of discriminative effects and inaccuracy of sample volumes.7,8 Improved heating of the injector head, improving elution from the rear of the needle, was one response to it;9 introduction of the fast autosampler by Hewlett-Packard10 was the other. Fast injection combined with a cool injector head suppresses vaporization in the needle. It was not properly recognized, however, that this also changes the basic conditions for sample evaporation in the injector. Ever since there are the two modes of sample introduction schematically shown in Figure 1. Thermospray Injection. Thermospray results from partial solvent evaporation inside the needle. Without really knowing, thermospray injection was the technique always used by manual and classical autosampler injection of solutions in volatile solvents. Passage through the hot syringe needle causes some solvent to evaporate along the wall. Since the mechanical displacement is fast, the vapor builds up pressure. The liquid is overheated and expelled through the center of the needle. Violent evaporation upon depressurization causes the exiting liquid to explode and be nebulized near the needle exit. The resulting small droplets initially move at a high velocity (exceeding 10 m/s), but friction rapidly slows them to the speed of the gas. The latter is about 2 mm/s in splitless injection and 80 mm/s when the split flow rate is 60 mL/min (4-mm-i.d. liner). If the distance to the column entrance is 40 mm, there is 500 ms to 40 s of time for evaporation. As shown in ref 3, 200-500 ms is sufficient for transferring the heat for solvent evaporation and heating of the sample if cooling of the chamber by several tens of degrees is accepted and a gas of high thermal conductivity is used (such as hydrogen). In fact, fluorescence mostly disappeared some 5-20 mm below the needle tip, suggesting complete solvent evaporation within this distance. (7) Grob, K.; Neukom, H. P. J. High Resolut. Chromatogr., Chromatogr. Commun. 1979, 2, 15-21. (8) Grob, K.; Rennhard, S. J. High Resolut. Chromatogr., Chromatogr. Commun. 1980, 3, 627-634. (9) Grob, K.; Neukom, H. P. J. Chromatogr. 1980, 198, 64-69. (10) Snyder, W. D. Technical Paper 108, Hewlett-Packard, Palo Alto, CA, 1985.

Figure 1. Sample liquid leaving the syringe needle: thermospray after partial solvent evaporation inside the syringe needle versus band formation when evaporation in the needle is suppressed, e.g., by fast autosampler injection.

If the old term “flash evaporation” suits a vaporizing technique, it is appropriate here. Thermospray injection results in vaporization from droplets and microparticles suspended in the gas phase. In fact, the dense liquid phase (fluorescence) turned into a fine fog some 0.5 mm from the needle exit. After 80 ms (two frames of the video), the fog was homogeneously spread over the chamber and had lost its impulse of movement. Then the gas slowly discharged it, seemingly more by dilution than by pluglike transfer.4 No fluorescence (perylene) remained on the glass wall, confirming that evaporation occurred from droplets and particles in the gas phase. Contact with the liner wall presupposes diffusion, which is particularly slow for the droplets or particles from which highboiling components are vaporized. Evaporation from particles rules out adsorption or degradation on active sites of the liner, that is, it is the most gentle process we can think of. From this description, thermospray seems poorly suited for samples containing large amounts of nonevaporating byproducts: if these byproducts are suspended in the gas as particles, they enter the column together with the solutes and severely contaminate its inlet. Practical experience taught us, however, that most of the contaminants are, nevertheless, deposited onto the liner wall, visible as a brownish dark ring. This is, in fact, the reason splitless injection (traditionally performed with thermospray) was always considered as the method of choice for matrixloaded samples. The visual experiments also provided some hints to resolve this contradiction to what was stated above for solutes: after injection of a solution containing 5% edible oil into a 5-mmi.d. tube, most of the microdroplets were transferred to the wall in less than 50 ms. It was speculated that electric charges formed during thermospray generated the necessary strong forces. It remains a key point to be explored which droplets or particles are attracted to surfaces and which others are not. Analytical Chemistry, Vol. 74, No. 1, January 1, 2002

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Figure 2. Injection with band formation into a gooseneck liner (a 6-cm section being shown) heated at 200 °C in an oil bath: selected frames of the video representing 40 ms (starting time specified at bottom); no carrier gas flow, i.e., simulating splitless injection (from ref 4).

Injection with Band Formation. Fast autosamplers inject with a needle dwell time in the injector of some 200 ms. This prevents solvent evaporation inside the syringe needle provided the head of the injector is at a far lower temperature than the center. Lacking solvent vapor acting as a propellant causes the sample liquid to leave the needle as a band, as shown by Qian et al.11 This band moves through long heated tubes, passes bends, and even performs slalom around baffles, being repelled from the hot surfaces by a cushion of solvent vapor.12 Evaporation from the band is negligible since the velocity is so high (around 10 m/s) that a distance of, for example, 4 cm between the needle and the column is covered in a few milliseconds, which is too short for transferring a significant amount of heat. When a perylene solution was injected through a cool needle into an empty liner, the band hit the hot bottom surface of the chamber. Most liquid was rejected, followed by violent movements. The video pictures in Figure 2 show the course of events after injecting 5 µL of a hexane solution into a 4-mm-i.d. liner with a restriction at the bottom (1 in frame A; gooseneck liner, Restek, Bellefonte, PA). A piece of fused silica (2) marked the column inlet, classically positioned some 5 mm above the bottom of the chamber to keep it above a possible deposit of septum particles and other debris. The band of liquid (3 in frame B) hit the bottom and was whirled upward (4) some 5 cm high (frames B and C, frames 0-40 and 40-80 ms after injection). Some liquid was evaporated while flying through the chamber, but most dropped back to the bottom, often to be rejected again (frame D). Some 150 ms after injection, most of the liquid settled at the bottom, forming a ball nervously dancing above the hot surface. It seemed to glide down to the funnel-shaped bottom (frame E) and finally evaporated from the outer wall of the inserted capillary inlet (5 in frame F, 400 ms after injection). Perylene deposited on to the polyimide has no chance of entering the column. When the capillary was mounted lower, with its entrance below the orifice of the restriction, sometimes liquid was driven directly into it or past it toward the split outlet. (11) Qian, J.; Polymeropulos, C. E.; Ulisse, R. J. Chromatogr. 1992, 609, 269276. (12) Grob, K.; De Martin, M. J. High Resolut. Chromatogr. 1992, 15, 399-403.

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The band of liquid must be stopped above the column entrance to avoid contact with metallic surfaces and prevent that solute material ends up being deposited below the column entrance from where it can no longer enter the column. The liquid must be held in this position during the 0.2-5 s typically required for solvent evaporation. Hence, no empty liners should be used for fast autosampler injection. Sucking the liquid into a packing of low thermal mass is the first option, deactivated wool of glass, quartz, or fused silica most widely being used. The liquid arriving first cools the nearest fibers to the solvent boiling point and thus prepares the site for receiving the main portion of the sample. The liquid ends up hanging in the wool as a droplet. The region is cooled to the (pressurecorrected) solvent boiling point until solvent evaporation is completed. It is then heated during several seconds and releases the sample components fractionated according to volatility. The process resembles that of programmed temperature vaporizing (PTV) injection, with the difference that the temperature of the injector block is constant and just that of the evaporation zone varies. The second option is trapping the liquid between obstacles. Liners, such as that with the Jennings cup,13 were designed to improve the mixing of sample vapor with carrier gassnot for stopping a band of liquid. It is not surprising, therefore, that many of them do not respond to the requirements of injection with band formation: the liquid may pass through the obstacle or perform violent, uncontrolled movements above it. The ideal liner guides the liquid into a small chamber from where it can no longer escape. The laminar liner (Restek) approaches this design. The liquid passes along the liner wall to the bottom, invited to do so by a funnel-like entrance. It is trapped there because rapid vapor formation in the vicinity of hot surfaces hinders it entering the narrow spaces on the ways backward or forward. Solvent evaporation then occurs from one or a few droplets dancing on a hot surface. When solvent evaporation is complete, the solute material is deposited onto the surface and evaporates from there. (13) Jennings, W. G. J. Chromatogr. Sci. 1975, 13, 185-187.

Figure 3. Strategies for injection and sample evaporation in hot injectors.

OPTIMIZATION OF CONDITIONS Having these two evaporation processes in mind, optimization of conditions becomes more rational. The analyst must first decide whether to inject through a cool or a hot needle, hence to give preference to band formation or thermospray (Figure 3), taking into account the injection system available. Thermospray. The possibility of using empty liners is an advantage of thermospray injection, since this reduces the risks of adsorption, degradation, or retention on active or retaining (contaminated) surfaces. Thermospray enables the injection of extremely high boiling components at gentle conditions because the material may enter the column as aerosol rather than vapor. Packings and obstacles do not have a noticeable positive effect on evaporation, because nebulized material passes through them like fog or smoke is blown through the forest. Thermospray requires a hot needle wall. In fact, the optimum injector temperature might often primarily be determined by the needs of thermospray, i.e., solvent evaporation, rather than solute evaporation. The needle temperature should be at least some 100 °C above the solvent boiling point. A well-heated rear part of the needle is particularly important: (i) a high solvent vapor pressure built up there is most effective in expelling the liquid, (ii) it better overcomes the cooling by sample introduction, and (iii) it minimizes the losses of high-boiling solute material.9 The maximum temperature of the needle is determined by the injector parts surrounding it. The temperature regulated and given on the readout as “injector temperature” does not correspond to that of the upper part of the injector. Injectors for fast autosamplers have a particularly cool top. Hence, even when the inserted needle has reached the temperature of its environment, it may be far cooler than indicated and may not be sufficiently hot for thermospray (particularly when solutions are in higher boiling solvents). In fact, injectors designed for thermospray (e.g., that from CE Instruments/ThermoFinnigan) keep temperature nearly constant up to the septum.14 On the other hand, even very fast injection cannot suppress evaporation inside the needle if the top of the injector is heated near to the injector temperature. Hence, the injector is optimized either for nebulization (and hot needle injection) or for use with a fast autosampler. It is unreasonable to care for a high injector temperature in order to obtain efficient thermospray but inject too rapidly for the (14) Grob, K. Split and Splitless Injection in Capillary GC, 3rd ed.; Hu ¨ thig: Heidelberg, 1993; p 444.

needle to reach this temperature. It takes 3-5 s to heat the inserted needle, which leads to the “hot needle” technique:7 the sample is withdrawn from the needle into the barrel of the syringe, the needle is allowed to heat up in the injector during 3-5 s, and then the plunger is depressed as rapidly as possible. Hot needle injection originates from minimization of the problems resulting from evaporation inside the needle. Today we know that the advantages also have to do with optimized nebulization. Experience tells us that hot needle injection with long (e.g., 3 in., 71 mm) syringe needles provides more reproducible results. Now the explanation is at hand: it improves thermospray, because long needles reach further into the well-heated zone of the injector, the thermal capacity of the needle is higher (thick walls are preferable for the same reason), and the vapor pressure built up at the rear of the needle is higher, resulting in stronger overheating of the liquid. For splitless injection, the center of sample evaporation should be slightly above the column entrance, for 80mm liners anyway calling for a 71-mm needle. For split injection, a shorter needle would leave more room for mixing the vapor with carrier gas, but nevertheless, the longer needle often provided the better results.15 To act as an efficient propellant, the solvent should be volatile, consume a modest amount of heat for evaporation, and, perhaps, have a low surface tension. Using a 51-mm 26S-gauge needle heated to 180 °C, all solvents with boiling points up to 100 °C were nebulized (sample volumes tested, 5 µL). It should be reminded that the upper part of some injectors reaches this temperature only with difficulty, if at all, even when settings are extremely high. Band Formation. The sample liquid leaves the needle as a band in the following three situations: (1) injection by a fast autosampler through a cool septum cap; (2) when the needle is short (10-20 mm) and passes through a cool injector head, also slower (including manual) injection largely results in band formation;5 (3) samples in a high-boiling matrix (undiluted mixture or solution in a high-boiling solvent) always form bands. A 5-mm plug of rather loose deactivated wool is sufficient to reliably stop the liquid. Any additional wool merely enhances the risk of adsorption or degradation. An alternative is column packing material supported (but not covered) by wool. The packing material is easily stirred 5 mm deep by the thrust of the liquid and should, therefore, be some 10 mm deep. Frits are not suitable because the high thermal mass of the sintered particles causes the liquid to be rejected.6 Of the liners with obstacles tested, only the laminar and the minilaminar liner reliably stopped the liquid.6 The gooseneck liner prevents contact with the metal surface at the bottom of the injector, but it does not provide satisfactory control of evaporation as it does not hold the liquid at a defined site. If the sample is released as a band, a rather short needle is sufficient. The band travels to the stopper even over long distances; i.e., the center of evaporation is determined by the position of the packing or obstacle rather than by the needle length. In split injection, the stopper should be located just below the tip of the inserted needle in order to provide room for homogenizing the vapor across the liner. For splitless injection, (15) Grob, K.; Neukom, H. P.; Hilling, P. J. High Resolut. Chromatogr., Chromatogr. Commun. 1981, 4, 203-208.

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Figure 4. Reducing matrix effect. Nebulization of the calibration mixture produces a fog that is efficiently carried into the column (left), while the droplets formed from the matrix of the sample transfer the solutes to the liner wall, from where they evaporate with more difficulty (right).

the vaporizing chamber should be filled from the bottom to the top and the stopper positioned only slightly above the column. MATRIX EFFECTS “Matrix effects” may play an important role in the analysis of matrix-loaded (contaminated) samples and mean that the sample influences the analytical result. In particular, absolute or relative peak areas for a given amount of a compound depend on whether the latter is injected in pure solvent or a sample extract. Matrix effects easily cause systematic errors. They tend to be insidious since they are detectable only through specific testing. “Enhancing” and “reducing” matrix effects are distinguished. The enhancing effect produces a larger signal for the compound in the sample extract than in the standard solution (i.e., results tend to be excessively high), while with the reducing effect the peak area is reduced in the sample. The two effects have different backgrounds. Thermospray Injection. Reducing matrix effects are frequent in thermospray injection. Visual observations suggest the following mechanism. Clean solutions produce a stable fog that also carries high-boiling solutes into the column (Figure 4, left). When the same solution is loaded with a high concentration of high-boiling or involatile matrix material, the resulting droplets are pulled to the liner wall (right). Being cool as long as they contain solvent, the droplets carry the higher boiling solutes along and “glue” them to the wall. In splitless injection, peaks turn out too small if the solutes are not evaporated from the layer of contaminants and transferred into the column during the splitless period. The reducing matrix effects were investigated using a silicone oil to imitate a nonvolatile matrix.16 With 5% oil in the sample, the peak of a component as volatile as n-decane was 15% too small and the areas of n-octadecane and higher alkanes were diminished (16) Grob K.; Bossard, M. J. Chromatogr. 1984, 294, 65-75.

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by 30-40%. Experiments with edible oil as matrix material and sterols as solutes showed even stronger deviations. These matrix effects disappeared when PTV splitless injection was used, presumably because cold injection avoids nebulization.17 For solutes of up to fairly high boiling points, reducing matrix effects can be avoided by injection directly into a dense plug of glass wool.18 In fact, in visual experiments, little fog left the plug at the bottom,5 indicating that most of the material was transferred to the wool and the sample evaporated from there (samples nebulized above the wool pass through it). The vicinity of the wool to the needle tip seems to suppress nebulization. Unfortunately, with this transfer to surfaces also the advantages of thermospray injection are lost; i.e., the problem is solved from the wrong end: the calibration with a clean mixture now performs as poorly as the analysis of the sample, rather than that the performance for the sample is enhanced to that of the calibration. Band Formation. The term “matrix-induced chromatographic response enhancement” was coined by Erney et al.19 and refers to a phenomenon well described for the analysis of pesticide residues in foods: peak areas per amount injected are larger in extracts of samples than in the matrix-free calibration mixture. Since peaks cannot represent more than 100% of the material injected, the smaller areas obtained for the calibration must be the problem (the opposite of the reducing matrix effects). Mu¨ller and Stan20 showed how peak areas developed during a series of 100 injections, a first 15 of a clean mixture of standards followed by 85 spiked extracts of spinach. The matrix material introduced by the first injections of the spinach extract caused the areas of critical components to increase. Then a plateau was reached, and later areas shrunk again, probably because of an excessive accumulation of contaminants. Enhancing matrix effects were primarily observed with packed liners, often (but not exclusively) in conjunction with band formation, i.e., deposition of the sample liquid onto surfaces. They are the result of a temporary deactivation of these surfaces by sample material: contaminants (which may be as volatile as water) deactivate the surface for some time and improve the conditions for the evaporation of the solutes, but largely disappear (evaporate or degrade) before the next sample is injected. Their source is insufficient deactivation: sample components obviously deactivate more efficiently than the treatment applied by the manufacturers. “Priming” of packed columns was based on the same principle: a sample was injected several times in short intervals before the analyses were started. Enhancing matrix effects are also observed in PTV and (to a smaller extent) on-column injection or for columns. They presuppose intense contact with surfaces, as obtained after injection with band formation and deposition of the liquid onto cooled surfaces. EVALUATION OF THE TWO APPROACHES The reader probably long started comparing the two ways of sample evaporation in order to find the preferable technique for his application. He should keep in mind that present instrumenta(17) Grob, K.; La¨ubli, T.; Brechbu ¨ hler, B. J. High Resolut. Chromatogr., Chromatogr. Commun. 1988, 11, 462-470. (18) Grob K.; Neukom, H. P. Chromatographia 1984, 18, 517-519. (19) Erney, D. R.; Gillespie, A. M.; Gilvydis, D. M.; Poole, C. F. J. Chromatogr., A 1993, 638, 57-63. (20) Mu ¨ ller, H.-M.; Stan, H.-J. J. High Resolut. Chromatogr. 1990, 13, 697-701.

tion is optimized either for thermospray or band formation (or neither of both). Hence, the choice is limited once he has bought the instrument. He should consider the following aspects: (1) Evaporation in the needle (thermospray) causes more sample to be injected than read on the barrel of the syringe as well as selective loss (discrimination) of high boilers. This was the reason for introducing fast autosampler injection, although with well-heated injectors discrimination can also be kept low. (2) Nebulized, fairly clean samples evaporate from droplets or particles suspended in the gas, thus from their own matrix. This is why hot needle injection tends to perform better than injection depositing the sample onto deactivated wool, even though it has classically been performed with raw glass liners (no specific data available). Thermospray is, therefore, preferable for the analysis of adsorptive, labile, and high-boiling solutes. This no longer holds true when the samples contain high concentrations of high-boiling or involatile components (large particles are attracted to the liner wall). (3) Both techniques may suffer from matrix effects. Enhancing effects are observed after deposition of the solute material onto surfaces and are primarily a problem for adsorptive and labile components. Thermospray injection may suffer from matrix material gluing solutes to the liner wall, which affects all higher boiling solutes. Solute evaporation is not more hindered than after deposition onto a surface by injection with band formation, but in comparison with the better performance of the clean solution, the losses are more obvious. (4) In principle, evaporation from a surface after injection with band formation produces clean vapor, leaving behind in the injector all the nonevaporating material, while thermospray also nebulizes involatile material. However, experience taught us that differences are not as large: column contamination after injection resulting in band formation is not always negligible, whereas that by thermospray injection is not a severe problem. (5) The accuracy obtained with split injection depends on the stability of the split ratio and on whether all solutes are split by the same ratio.21 Discrimination from a fluctuating split ratio results when different components arrive at the split point at different times (different split ratios). Evaporation from an initially cooled surface, following injection with band formation, is probably the most important fractionation mechanism (although under some conditions the split ratio may have restabilized before the solutes evaporate). Thermospray injection produces a vapor cloud with a homogeneous sample composition; i.e., a fluctuating split ratio does not result in discrimination. Another question is whether vapor and aerosol particles are split by the same ratio, which could be a different source of discrimination effects. (6) If fast autosamplers are used, sample evaporation in the injector fundamentally differs from that following manual injection. Hence, a method validated for fast autosampler injection cannot be carried out with manual injection or a slower autosampler. If thermospray is applied, manual and automatic hot needle injection perform equally. CONSEQUENCES FOR THE INJECTOR DESIGN With the concepts of sample evaporation in mind, we should be able to improve the design of the split/splitless injector. The (21) Grob, K. Split and Splitless Injection in Capillary GC, 3rd ed.; Hu ¨ thig: Heidelberg, 1993; pp 126-159.

basic design of the injectors presently in use is from the 1970s,22 i.e., from a time long before the fast autosampler was introduced. The thermospray mechanism was not understood at that time, but something similar was assumed since the sample was thought to evaporate just below the needle exit. Hence, the geometry of our injectors largely reflects the philosophy of thermosprayseven when used with fast autosamplers. The determining factors were as follows: (1) If the injector is designed according to the needs of splitless injection, it is also suitable for split injection, but not vice versa. (2) The volume of the vaporizing chamber must be sufficiently large for the intermediate storage of the sample vapors (splitless injection and split injection with low split flow rates). Two microliters of solutions in hexane and water (two extremes) produces some 300 and 3000 µL of vapor, respectively. (3) The internal diameter of the chamber cannot exceed 4-5 mm since the gas velocity during splitless sample transfer becomes too low otherwise.23 Furthermore, a larger bore increases the capacity less than proportionally to the volume because of increased mixing (dilution) with carrier gas (the vapors form something like smoke trails through the gas in the injector instead of displacing the gas similar to a piston). (4) To fill the chamber from the bottom (column entrance) to the top, the syringe needle must be sufficiently long to position the center of sample evaporation slightly above the column entrance. Since syringe needles longer than 3 in. (71 mm) are awkward to use, chambers of 80-mm length became standard. (5) The accessible volumes in the split outlet line must be small in order to prevent that pressure increase during solvent evaporation pushes sample vapor into this outlet. The following elements were introduced after 1978: (1) Improved heating of the injector head decreased discrimination against high-boiling solutes for hot needle injection.9 (2) The fast autosampler avoided sample evaporation inside the syringe needle.10 (3) The programmable inlet pressure (Wylie et al.24) enabled the increase of pressure during splitless injection (“pressure pulse”), which compresses the vapor cloud and thus enhances the capacity of the vaporizing chamber. At the same time, it accelerates the sample transfer into the column, allowing the use of wider bore chambers. It has been known for a long time that the internal volume of a 80 × 4 mm i.d. chamber (1 mL) is often too small to reliably hold the sample vapor.25,26 At modest inlet pressures it does not even house the vapor of 1 µL of a solution in methanol. Overloading is frequent and serious effects on quantitative analysis have been reported (e.g., refs 27 and 28). Elongation of the liner is the only efficient means of enhancing the injector capacity. For instance, a 16 × 4 mm i.d. liner would offer 2 mL of room. Reduced (22) Grob, K.; Grob, K. J. High Resolut. Chromatogr., Chromatogr. Commun. 1978, 1, 57-63. (23) Grob, K. Split and Splitless Injection in Capillary GC, 3rd ed.; Hu ¨ thig: Heidelberg, 1993; pp 273-287. (24) Wylie, Ph. L.; Phillips, R. J.; Klein, K. J.; Thompson, M. Q.; Hermann, B. W. J. High Resolut. Chromatogr. 1991, 14, 649-654. (25) Grob, K.; Biedermann, M. J. High Resolut. Chromatogr. 1989, 12, 89-96. (26) Hinshaw, J. V. J. High Resolut. Chromatogr. 1993, 16, 247-153. (27) Lee, H. B.; Szawiola, R.; Chau, R. S. Y. J. Assoc. Off. Anal. Chem. 1987, 70, 929-935. (28) Grob, K. J. Assoc. Off. Anal. Chem. 1988, 71, 76A-77A.

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far backward toward the top, while the higher boiling solutes evaporate last and form a small vapor cloud staying near the column entrance. Their transfer into the column is correspondingly fast. This corresponds to the process used for large-volume splitless injection by the overflow technique.29 Since the upper part of the chamber primarily serves for intermediate storage of solvent and volatile compounds, its temperature may be rather low, just sufficient to prevent the band touching the liner surface. For split injection, the band stopper is placed higher in order to leave enough room for spreading the vapor across the liner.

Figure 5. Ideas for an improved design of an injector for split/ splitless injection with thermospray or band formation.

mixing with carrier gas increases capacity more than proportionally to the volume and should enable injection of about 2 µL of methanol (1.5 mL of vapor at 30 kPa and 250 °C), 1 µL of water (1500 µL), 3.5 µL of dichloromethane (1600 µL), or 6 µL of hexane (1550 µL). With an inlet pressure of 160 kPa, the capacity is approximately doubled. Such elongation of the chamber presupposes, however, a new concept on how to bring the center of evaporation so far down the chamber to be situated just slightly above the column inlet. The options depend on whether injection is performed with thermospray or band formation. Optimized Thermospray Injector. An optimized thermospray injector should comprise a nebulizer of small internal volume releasing the sample near the column entrance. It could consist of the syringe needle or a separate device. The thermal mass and thermal conductivity of the material should be such that there is no strong cooling by solvent evaporation. The nubulizer could be positioned laterally near the bottom of the chamber (Figure 5), which probably presupposes a liner made of deactivated steel, or enter at the bottom parallel to the column. Injection with Band Formation. Solutions in volatile solvents injected with band formation can be shot through long hot tubes without significant evaporation, since vapor cushions prevent the transfer of the liquid to the liner wall. A short needle, merely entering the liner by a few millimeters (totally 15-20 mm long), is sufficient. If the top of the injector is cool, there might not even be a need for a particularly fast injection. The needle tip must be such as to rule out mechanical spray. For splitless injection, the band is stopped at the bottom of the chamber by means of a packed bed or a trap. As the solvent and the highly volatile components evaporate first, they expand

16

Analytical Chemistry, Vol. 74, No. 1, January 1, 2002

OUTLOOK It seems odd that a key technique of capillary GC, such as hot split and splitless injection, still needs basic development work decades after its introduction and with maybe 200 000 analysts using it for daily routine. Improvement is needed, because the processes are not under a control satisfying the requirements for a professional tool, and all too often poor quantitative results are obtained for reasons that cannot be put aside blaming the analyst. Often methods do not tell what conditions to select (fast autosampler/manual injection? needle length? side port hole in the needle? hot needle injection? liner dimension? packing? position of the packing? etc.), leaving this “up to the responsibility of the analyst”, as it may be put to avoid saying that nobody could tell him. Many methods describe in great detail how the round flask should be rinsed (rather an insult to experienced analysts), but then provide hardly any advice on the often most important source of error. The distinction between thermospray and band formation should help understand phenomena and optimize conditions in a rational manner. Methods should specify whether they were validated for thermospray or band formation. Basically they should be validated for both, since the two techniques run on different types of instruments and users commonly just have access to one of them. In the long run, the question needs to be answered of whether vaporizing injection could be standardized on a single principle of sample evaporation. This would simplify the specification of methods but also deprive us of the possibility to choose the other technique because of better performance for a particular sample. In fact, it is not obvious which technique to prefer. Insights resulting from the videos on the evaporation process make us believe that substantially improved injectors could be designed, improving the quality of the results, enabling us to inject somewhat larger volumes, and helping us to better master the critical conditions. Received for review July 6, 2001. Accepted October 12, 2001. AC0107554 (29) Grob, K.; Brem, S.; Fro ¨hlich, D. J. High Resolut. Chromatogr. 1992, 15, 659-664.