MASS SPECTROMETRY
4.1. Gas Chromatography–Mass Spectrometry Interfacing
For use in combination with packed-column GC, a variety of interfaces for GC–
MS were developed. The aim of these devices was to achieve analyte enrichment,
Table 6 Comparison of Some Features of Various Mass Analyses Double-focusing Linear
Feature sector quadrupole Ion trap TOF
Scanning Fast Fast Super fast
Vacuum Critical Bath gas
Mass range 10,000 4000 2000 No limits
Acquisition Full-scan, SIM Full-scan, SIM Full-scan, SIM Full-scan
Resolution High Unit-mass Unit-mass High
Mass accuracy Accurate Nominal Nominal Accurate
(1 ppm) (5 ppm)
Principles and Instrumentation 19
Figure 5 Schematic diagram of the open split coupling for GC–MS.
i.e., a better ratio between analyte and carrier gas. The jet separator, the Watson- Biemann or effusion interface, and the membrane interface are the most fre- quently applied devices of the type.
With the introduction of open capillary columns for GC and GC–MS, an analyte enrichment interface is no longer required as the optimum flow rate of such a column is readily amenable to the vacuum system of a benchtop GC–MS system. At present, two types of GC–MS coupling are applied, i.e., the direct coupling and the open split interface. In the direct coupling, the column effluent of the GC column is directly introduced into the ion source of the mass spectrome- ter. While this approach is very simple, it has some disadvantages that are avoided by the use of an open split coupling (Fig. 5). In a direct-coupled GC–MS, the column outlet is at high vacuum, resulting in changes in the chromatogram similar to those obtained from a GC–FID. As the complete output of the GC column is introduced into the source, the risk of source detuning and contamination is higher, e.g., due to the solvent pulse, flow-rate changes during temperature pro- gramming, and sample contaminants. While in direct coupling, the vacuum sys- tem must be switched off for changing the GC column; this is not required with the open split coupling.
4.2. Sample Pretreatment and Large-Volume Injection
A major disadvantage of the small column internal diameters used in open capil- lary GC is the limited injection volume. This limitation can partly be reduced by the use of preconcentrating sample pretreatment, such liquid-liquid extraction (LLE), solid-phase extraction (SPE), and solid-phase microextraction (SPME).
These sample pretreatment techniques are widely applied throughout the daily practice of GC–MS, as exemplified by the sample pretreatment procedures de- scribed in the various chapters of this book.
In addition, large-volume sample introduction techniques have been intro- duced for open capillary GC, e.g., on-column injection, loop-type injection, and
20Niessen
Table 7 Large-volume Introduction Techniques for Capillary GC ON-COLUMN INJECTION
Two types of evaporation techniques can be applied:
Conventional retention gap technique
Sample is injected at a temperature below the solvent boiling point. If the retention gap can be wetted by the solvent, a flooded zone is formed. The solvent film evaporates from the rear to the front and volatile analytes are reconcentrated by the solvent trapping effect. In addition, phase soaking effects reconcentration of the analytes due to the increased retention power of the thicker stationary phase. Less volatile components remain spread over the retention gap and are reconcentrated by the phase-ratio-focusing effect.
Partially concurrent solvent evaporation (PCSE)
Sample is injected into the GC under conditions that cause the major part of the solvent to evaporate while the remaining solvent floods the retention gap; that is, the solvent introduction rate is higher than the evaporation rate. In this way, about 90% of the introduced sol- vent can be evaporated during introduction. Volatile analytes are reconcentrated due to phase soaking and solvent trapping in the re- maining solvent film. Less volatile components remain spread over the retention gap and are reconcentrated by the phase-ratio- focusing effect.
LOOP-TYPE INJECTION
The sample in the loop is pushed into the GC by the carrier gas. Two types of solvent evaporation technique can be applied:
Fully concurrent solvent evaporation technique (FCSE)
Sample is injected at a temperature above the solvent boiling point. The sample is completely evaporated during injection. No flooded zone is formed. Volatile analytes co-evaporate with the solvent. Less volatile components remain spread over the retention gap and are reconcentrated by the phase-ratio-focusing effect.
Co-solvent trapping
A small amount of a higher boiling co-solvent (e.g. octadecane) is added to the main solvent to create a layer of condensed liquid ahead of the main evaporation site. The main solvent evaporates concurrently, and part of the co-solvent evaporates together with the main solvent.
Boiling point and amount of co-solvent must be adjusted such that some co-solvent is left behind as a liquid and spreads into the retention gap. Volatile analytes are reconcentrated due to solvent trapping in the co-solvent. Less volatile components remain spread over the retention gap and are reconcentrated by the phase-ratio-focusing effect.
PrinciplesandInstrumentation21 PTV INJECTION
A programmed temperature vaporizer injector (PTV) basically is a split-splitless injector with temperature control, i.e., the vaporizer chamber can be heated or cooled rapidly. Three types of large-volume introduction techniques can be distinguished:
PTV solvent split injection
The sample is injected in a packed liner with an open split exit at an injector temperature below the solvent boiling point. Volatile compounds co-evaporating with the solvent are lost. After solvent evaporation, the analytes retained in the liner are transferred to the GC column. The maximum introduction volume which can be injected ‘‘at once’’ mainly depends on the liner dimensions: a 1 mm i.d. liner can hold 20–
30àl of liquid, 3–4 mm i.d. liners can hold up to 150àl of liquid. Higher sample volumes have to be introduced in a speed-controlled manner where the speed is adjusted to the evaporation rate.
PTV large-volume splitless injection
The sample is introduced in a packed liner at a temperature below or close to the solvent boiling point. The split exit is kept closed, i.e., the flow rate through the liner is equal to the column flow rate. The evaporating solvent is vented via the GC column. Volatile components co-evaporating with the solvent are trapped in the swollen stationary phase of the GC column.
PTV vapor overflow
The sample is rapidly injected into a packed liner at a temperature far above the boiling point of the solvent. During solvent evaporation, the split exit is closed but the septum purge is wide open; the evaporating solvent escapes through the purge exit. After solvent evaporation, the injector is heated to effect transfer of the analytes to the GC column. The technique has also been carried out in a conventional split/
splitless injector.
Source: Ref. 21.
22 Niessen
programmed temperature vaporizer (PTV) injection. These techniques enable in- jection of volumes as large as 100 àl onto the GC column. These techniques have recently been reviewed by Hankemeier [21] and are briefly described in Table 7. In on-column and loop-type large-volume injection systems, a retention gap is used. This is an uncoated deactivated fused-silica injection column that enables the reconcentration of broadened bands. The performance of both on- column and loop-type injections can be further enhanced by the use of a solvent vapor exit (SVE). The SVE is a solvent release system that helps to protect the GC detector from vapor and to accelerate solvent evaporation. The SVE is posi- tioned prior to the GC column [21].
4.3. Analyte Derivatization
Compounds that not amenable to GC analysis, either because of limited thermal stability or insufficient volatility, can sometimes be made amenable by means of derivatization. A general aim in this type of derivatization is the reduction of analyte polarity by chemical substitution of active protons in the analyte. In MS, derivatization may result in additional effects, e.g., enhancement of the intensity of the molecular ion, changes in the fragmentation directing functionality, and/
or improvement of the ionization efficiency. A clear example of the last is the introduction by derivatization of fluorine groups in a molecule, enhancing its amenability to electron-capture detection. A wide variety of derivatization re- agents are available [22]. An overview of frequently applied derivatization agents for various compound classes is given in Table 8.
Table 8 Common Derivatization Reagents for GC–MS
Increase in Mr
Compound class Group replacing per derivatized
(functional group) Reagent functional group functional group
Alcohols (EOH) TMS EOSi(CH3)3 72
TBDMS EOSi(CH3)2C(CH3)3 114
Acetylation EOC(CO)CH3 42
Carboxylic Acids Methylation EOCH3 14
(EOH) TMS EOSi(CH3)3 72
Amines and Amides Acetylation ENRC(CO)CH3 42
(ENRH) TMS ENRSi(CH3)3 72
TFA ENRC(CO)CF3 96
Carbonyl Compounds Methoxime ECCNEOCH3 29
(CCO) Oxime/TMS ECCNEOSi(CH3)3 87
Phenylhydrazone ECCHENHEC6H5 102
Principles and Instrumentation 23
Derivatization obviously complicates the analytical method, as a (series of) time-consuming step(s) must be included. Sometimes, the derivatization gener- ates additional problems due to artifact formation. Routine derivatization in trace quantitative analysis is often difficult to perform.
4.4. Data Acquisition and Processing
Two general modes of data acquisition are available in MS: full-scan acquisition and selective ion monitoring (SIM). In full-scan analysis, a continuous series of mass spectra is acquired during the chromatographic run. For high-efficiency open capillary GC columns, sufficiently fast scanning is required in order to ac- quire a sufficient number of data points (typically 10 to 20) to adequately describe the chromatographic peak profile. However, in routine quantitative analysis of a limited number of components, better results in terms of lower detection limits are achieved by the use of SIM, in which the intensity of a (number of) ion(s) is monitored. The choice between full-scan and SIM acquisition in a particular application depends on the required detection limit and information content.
As a result of data acquisition, a three-dimensional data array along the axes time, m/z, and ion intensity is generated in the data system. This array can be processed in a number of ways. In the total-ion chromatogram (TIC), the total ion intensity per spectrum is plotted as a function of time. This provides more- or-less universal detection, with a chromatogram comparable to a FID chromato- gram. From the peaks in the TIC, a mass spectrum may be obtained. In order to minimize concentration effects on the spectrum quality in narrow GC peaks, an averaged mass spectrum is often obtained. Background subtraction can also be applied to enhance the spectrum quality. The mass spectrum may be computer searched against a library of mass spectra to enable provisional identification (see section 5.1). An alternative to the TIC is the mass chromatogram or extracted ion current (XIC) chromatogram, where the ion intensity of a selected m/z is plotted as a function of time.
In addition to these general modes of data processing, a variety of more specialized procedures are available, e.g., for quantitative analysis, including fully automated peak integration and calibration by linear regression.