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Anal. Chem. 1989, 61,58-65
Characterization and Optimization of Flame Photometric Detection in Supercritical Fluid Chromatography Susan V. Olesik,* Lars A. Pekay, a n d Elizabeth A. Paliwoda
Department of Chemistry, The Ohio State University, 120 West 18th Avenue, Columbus, Ohio 43210
The flame photometrlc detector (FPD) was optimized for the anaiysls of organosulfur compounds by supercrttlcal fluid chromatography (SFC). The flame gases used In thls study were hydrogen and oxygen. The optimum flow rates for hydrogen and oxygen were determined. The effect of placing the pressure restrictor dmerent distances away from the flame was examined. The flame chemisky that resunb when carbon dloxide flows Into a hydrogen-rich flame was characterized. The coelutlon of hydrocarbons wlth sulfur-containing compounds did not markedly quench the sulfur dimer signal for concentrations that were approprlate for capillary SFC. Detection limits as low as 0.1 ng of sulfur were achieved with detectivities of 8 pg/s.
In 1962 a German patent was issued to H.Draeger and B. Draeger for the determination of sulfur and phosphorus using a hydrogen-rich flame. Soon thereafter Brody and Chaney applied this technique to the detection of gas chromatographic effluents ( 1 ) . Due to its high sensitivity and high selectivity, flame photometric detection in gas chromatographic analyses is now the method of choice for analysis of compounds containing sulfur and phosphorus. The capability of combining an efficient separation process with a sulfur-specific detector for high molecular weight species is highly desired in many areas of chemistry, such as clinical analysis, petroleum analyses, pharmaceuticals, and biotechnology. Unfortunately, the flame photometric detector is not easily interfaced to high-performance liquid chromatography (HPLC) which is the most common separation technique for high molecular weight species. Difficulties found with HPLC/FPD (flame photometric detection) include high detection limits, excessive quenching by the presence of organic molecules, and differential volatilization of solutes and solvents. Accordingly, sample concentration methods followed by analysis with atomic fluorescence or atomic absorption spectroscopy are often necessary to quantify the sulfur content in nonvolatile species. One notable exception to this arduous task should be pointed out. McGuffin and Novotny ( 2 ) showed that low detection limmts could be achieved for nonvolatile samples when detecting phosphorus by pHPLC/dual-flame FPD. No information on the possible detection of sulfur by this technique is available to date. Supercritical fluid chromatography (SFC) is used with increased frequency for the separation of high molecular weight species. In addition, supercritical fluid chromatography is much more easily interfaced to information-intensive techniques such as Fourier transform infrared (FT-IR) ( 3 , 4 )and mass spectrometry (5). Recently, flame photometric detection in SFC was demonstrated (6),but the detection limits for sulfur were high. In addition, other concerns such as variation of base line with pressure were cited as a problem. We describe in this paper the characterization and optimization of SFC/FPD using hydrogen and oxygen as flame gases and supercritical carbon dioxide as the carrier gas. This paper shows that after optimization, SFC/FPD is a viable technique for total sulfur detection of nonvolatile compounds.
EXPERIMENTAL SECTION The instrumentation used in this study is shown in Figure 1. The mobile phase was supercritical fluid grade carbon dioxide (Scott Specialty Gases). Carbon dioxide was pressurized by an ISCO micro-LC500 syringe pump (ISCO Nebraska) and then pumped through a heated interface of original design to a Valco W-series high-pressureinjection valve that was fitted with a 60-nL rotor (Valco Instruments, Houston, TX). The heated interface was built and installed to allow for heated injections and/or constant temperature transfer of fluids from a supercritical fluid extraction system. The solvent and solutes were then delivered to a 100-wm i.d. fused silica column (J&W SFC series DB-1701), which was maintained in a Hewlett-Packard HP5890A gas chromatographic oven at a temperature above the critical temperature of carbon dioxide (32 "C). The detector that was optimized in this study was the standard Hewlett-Packard flame photometric detector (Model 19256A). No mechanical changes were made in the detector design, but very significant changes were made in the flame makeup (as described below) to achieve optimum performance. The filter used to monitor the Sz chemiluminescence was a broad band-pass filter ( A i = 10 nm) with peak transmission at 394 nm. A chromatographic column was not necessary for every experiment described. In some cases the column was replaced with a 250-wm fused silica open tube (PolymicroTechnologies, Inc.). The restrictor used for most experiments was a 15-fimi.d. fused silica tube, but integral restrictors and Lee restrictors (Lee Scientific)also were found to perform well with this detector. The connectionbetween the column and the restrictor was made with a Valco zero dead volume union. The analog signal from the FPD was monitored by both a chart recorder and a Zenith 2200 (IBM-ATcompatible)that was equipped with a Data Translations DT282112-bit,50-kHz analog to digital (A/D) board. Integration and data analysis were accomplished by using original programs written in the ASYST Laboratory analysis programming environment. The fuel and oxidant gases used in the FPD were hydrogen and oxygen (Instrument grade, Matheson gases). The model compounds used in this study were methyl p-tolyl sulfide, thiophenol, benzo[b]thiophene,phenyl sulfide, and dibenzothiophene. Each compound was used as delivered by Aldrich. The solvent used to inject the samples onto the column was anhydrous spectrophotometric grade methyl alcohol (Mallinckrodt Chemicals). RESULTS AND DISCUSSION Mechanism. In the flame photometric detector the chemiluminescent emission of the excited Sz molecule is monitored to quantify the sulfur content in a sample. Sulfur species that are present when sulfur-containing compounds are burnt in a hydrogen-rich flame include HzS,SOz, SO, SH, Sz, and CS. Since the dissociation energies of these compounds are very similar, the concentration of species in the flame is greatly dependent on the temperature and composition of the flame. The reaction sequence that occurs upon addition of a sulfur-containing compound to a hydrogen flame that is sustained by oxygen is sulfur compound
-+
HzS
+ H 3 SH Hz SH + S Sz + H
H2S
0003-2700/89/0361-0058$01.50/00 1988 American Chemical Society
2
(1) (2)
(3)
ANALYTICAL CHEMISTRY, VOL. 61, NO. 1, JANUARY 1, 1989
50
! 50 -
40 -
PUMP
COLUMN
INTERFACE
30
20
-
Figure 1. SFC/FPD instrumentation.
Reaction 3 is the rate-determining step of this sequence (7). Subsequently, although not unambiguously proven, the reaction that probably causes the electronic excitation of the sulfur dimer is reaction 4 (8,9). The emission observed from
H
+ H + S2
-+
H2 + S2*
(4)
the excited S2 dimer corresponds to the transition from the first excited state B3Z,- to the X3Z[ ground state (9). The most intense bands in the Sz emission envelope are those at 384 and 394 nm (10). The FPD system that was optimized in this study selectively monitored the band at 394 nm. Another proposed mechanism for the production of excited sulfur dimer is shown in reactions 5 and 6 (11, 12). While
HS 2s
+ H + S + H2
+ M + S*2 + M
the sequence involving reactions 5 and 6 cannot be conclusively ruled out as a contributor to the sulfur dimer emission that is measured in H2/Ozflames, reaction 4 seems most probable. For example, Gaydon has shown that only the lower vibrational states of the B3Z[ sulfur dimer excited state are populated in Hz/Oz flames, which is expected if reaction 4 is the predominate cause of sulfur dimer excitation (9). Also, as shown later in this article, the quenching of the sulfur dimer emission due to the presence of hydrocarbons is best described by the disruption of reaction 4 rather than reactions 5 and 6. Optimization of Flame Gas Conditions. Due to the low temperature of a hydrogen-rich flame and its associated low heat capacity, the flame gas composition changes markedly with minor changes in flame gas flow rates. Therefore, considerable care must be taken in comparing the chemistry in various hydrogen-rich flame photometric detectors. For example, all experiments in this study used a hydrogen-rich Hz/Ozflame while most other studies of flame photometric detection of sulfur have used a hydrogen-rich H2/air flame. Much different reaction chemistry is found in these two flames (due to differences in flame temperature and quantity of radicals present in the flame) even though the reaction mechanism for the formation of sulfur dimer may very well be the same in each. For example, as described later, the presence of hydrocarbons affects the sulfur dimer emission much differently in this detector with Hz/02flame gases than is found in Hz/air flames. Accordingly, for comparative purposes the flame photometric detector with Hz/Oz flame gases was first optimized under standard gas chromatographic conditions. Standard operating conditions for this detector (as described by the manufacturer) involve initially mixing the carrier gas with a sheath of hydrogen. Subsequently, oxygen is not premixed with hydrogen but is brought to the flame jet through holes at its base. This is exactly opposite the flow configuration that was used in the original Brody-Chaney design in which the carrier was mixed with oxygen. Hydrogen and oxygen flow
io
-
0 1 . .
02 0
,
,
,
,
,
,
,
t
,
0 30
,
.
,
,
.
.
.
,
,
0 40
0,/ H ~ VOLUME UA no Figure 2. Effect of 02/H2 fiow-rate ratio on measured sulfur dimer emission in GC mode. Carrier gas was helium.
rates, as well as the detector temperature, were optimized with helium carrier gas using Simplex V (Statistical Programs, Houston, TX). Two simplex runs were undertaken. Both runs had different starting values to ensure the validity of the optimum values achieved. The optimum conditions were determined to be an oxygen flow rate of 24 mL/min, and hydrogen flow rate of 70 mL/min which corresponds to an 0 2 / H 2ratio of 0.34 for a helium flow rate of 20 mL/min (Figure 2). These values compare well with the previous work (18-40 mL/min O2and 80-200 mL/min H2 for an Oz/Hzflow rate ratio = 0.24.3) obtained by methods other than statistical design (13). The detector temperature was found to be statistically insignificant in the range of 175-300 "C. Simplex optimization (14)and factorial design (15) were used to optimize the gas flow rates for the SFC/FPD system. These data, combined with a complete matrix experiment, were used to provide the response surfaces shown in Figure 3. The detector temperature was also found to be statistically insignificant over the range of 175-300 "C. Figure 3A is a response surface showing the dependence of sulfur emission (signal to noise (S/N) values that were corrected for background emission) on oxygen and hydrogen flow rate. The optimum conditions for sulfur detection in SFC mode were determined to be 239 mL/min H2 and 45 mL/min O2 which correspond to an Oz/H2= 0.19 with a carbon dioxide volumetric flow rate of 32.0 pL/min. These conditions provide maximum S/N for the sulfur dimer emission and minimum background (Figure 3B). One important feature of the background dependence on flame gas composition is that over a very broad range of hydrogen and oxygen flow rates the background is flat. The background emission is probably primarily due to carbon dioxide emission. In carbon monoxide flames (16) and carbon dioxide afterglow (9) narrow emission bands superimposed on a continuum are found in the visible spectrum that are assigned to carbon dioxide (17). The carbon dioxide emission bands are believed to correspond to the transition of bent carbon dioxide excited state lB2to the linear ground state x1 E+, (16),whereas the continuum in carbon monoxide flames has been described as being due to the chemiluminescent formation of excited carbon dioxide by reaction of carbon monoxide and oxygen (9). Previous studies (9) also showed that the two sources of emission behaved differently when the flame temperature was varied. As the flame temperature was lowered the continuum signal decreased drastically while the intensity of the emission bands was invariant.
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ANALYTICAL CHEMISTRY, VOL. 61, NO. 1, JANUARY 1, 1989 Ll
(without carrier gas), 2700 K (21); Tzis initial temperature of the carrier gas at the flame base, 523 K; T3is approximate thermodynamic temperature of the flame after the addition of carrier gas; C is the molar heat capacity of the flame, 12.4 cal/(mol K); Cp$s the heat capacity of the carrier gas at 2700 K, 4.98 cal/(mol K) for helium (22) and 14.2 cal/(mol K) for carbon dioxide (23); and X is the mole fraction of carrier gas in the flame, 0.18 for 20 mL/min helium and 0.13 for carbon dioxide flowing at a rate of 32 pL/min at 4000 psi and 40 OC. The heat capacity of the flame was determined from Cpl
Variatlon of (A) measured sulfurdimer emission and (e) background emission as a function of H2 and O2flow rates. Sample injected was 0.25 pg of benzo[b]thiophene. Figure 3.
Figure 3B shows that the intensity of the background quickly vanished as the relative proportion of hydrogen in the flame was increased, and under optimum conditions the background signal was negligible with only the dark current from the photomultiplier being measurable. Increasing the hydrogen content of the flame decreases the flame temperature and therefore the background observed in this configuration of the FPD is believed to be formed by a reactive process similar to that which formed the continuum in carbon monoxide flames. The most probable reaction for the formation of the carbon dioxide continuum emission in this work is the inelastic collision shown in reaction 7 which has been predicted theoretically and proven experimentally in reaction dynamics studies (18, 19). The formation of this collision complex causes facile transfer of energy between hydrogen radicals and carbon dioxide (18). Cop H C02-H COP hv H (7)
+
-+
-
+ +
An interesting comparison can be made between optimum conditions for gas chromatography and supercritical fluid chromatography using carbon dioxide. The optimum Oz/Hz ratio is lower in SFC/FPD than in GC/FPD. The temperature of the flame is strongly dependent on this ratio. As the 0 2 / H zratio decreases so does the flame temperature. In a chromatographic analysis the other variable that controls the temperature of the flame is the identity of the carrier gas. The variation in the thermodynamic flame temperature caused by injecting carbon dioxide carrier gas into the flame as opposed to injecting helium was compared. Assuming thermodynamic equilibrium is reached in the flame, the heat lost by the flame should equal the heat gained by the carrier gas. Hence the following equation can be used to calculate the change in temperature caused by carrier gas flow (20): where TI is approximate thermodynamictemperature of flame
= (XAC,A+ XBC,B ...)
(9)
where A, B, ..., C,, and X are the molecular and radical species that makeup the flame, their molar heat capacities, and the mole fraction, respectively. For hydrogen-oxygen flames with Hz/Oz flow rate ratios equal to approximately 0.2, the main components of the flame are water and hydrogen with approximate mole fractions of 0.4 and 0.6, respectively (24). The mole fractions of hydrogen, hydroxyl, and oxygen radicals present under these conditions are 0.04,0.008, and
