Anal. Chem. 1994,66, 1334-1338
Quantification of Compound Classes in Complex Mixtures and Fuels Using HPLC with Differential Refractive Index Detection Charles W. Sinkt and Dennis R. Hardy’ Naval Research Laboratory, Chemistry Division, Code 6 780, Washington, D.C. 20375-5342
The chemical composition of liquid hydrocarbon fuels that affect engine combustion performance has historically been defined by three principal classes: saturates, monocyclic aromatics, and dicyclic aromatics. These classes may be separatedby normal-phaseanalytical-scale high-performance liquid chromatography and are conveniently observed as the response of a differential refractive index (DRI) detector. Obtaining accurate quantitativeanalysis is not straightforward because calibration standards for determining the response factor for each compound class are not available. This problem arises because the refractive index of each class varies from fuel to fuel and is not generally known accurately enough for quantitative analysis. However, the linearity of the detector response with analyte refractive index for a fixed volume of sample makes it possible to determine the relative compound class composition as well as the refractive index of each class from the peak areas measured with a DRI detector at the time of separation by HPLC. This is accomplished by obtaining the chromatogramsof each sample using two different mobile phases of different refractive indexes, such as hexane and nonane in this study. The method has been applied to jet fuels in this paper, but the extension to other hydrocarbon fuels such as gasoline and diesel fuel is obviously possible. The fact that fuel composition can affect the performance of jet turbine engine combustion is well recognized. One compositional parameter, the total aromatics (a v/v), is defined for all commercial and military fuel specifications governing procurement. The current approved method for compound class quantitation, ASTM-D-1319-89, or the fluorescent indicator adsorption method (FIA), gives quantitative values for the saturates, olefins, and total aromatic hydrocarbons.‘ The FIA method, which was developed for gasolines and extended to jet fuels, uses dyes that are adsorbed by the different compound classes as they are separated on a silica column by displacement chromatography. Quantitation is accomplished by measuring the length of the column of the different fluorescing dyes under ultraviolet light. This technique is time consuming and is not generally applicable to all hydrocarbon fuels, especially diesel fuels, which exceed the boiling range limitations of the method. A review of other techniques that have been employed to obtain quantitative determinations of the compound classes in fuels is presented in a report by Ashraf-Khorassani et al.* that also details the + Current address: Department of Chemistry, Edinboro University of Pennsylvania, Edinboro, PA 16444. (1) Hydrocarbon Types in Liquid Petroleum Products by Fluorescent Indicator Adsorption, ASTM-D-1319-89; ASTM Vol. 05.03,Section 5 , 1991.
1334 Analytical Chemistry, Vol. 66, No. 8, April 15, 1994
use of supercritical fluid chromatography in obtaining quantitative analysis of hydrocarbon classes in various fuels. Previous attempts to use high-performance liquid chromatography with differential refractive index detection (HPLC/DRI)3-5 to obtain a separation similar to the FIA method and to quantify the compound classes have met with limited success. This is largely due to thedifficulty ofobtaining an appropriate detector response factor for the aromatic fraction(s). Most workers have tried to analyze the aromatic fraction as a single analyte and have not been able to obtain a detector response factor that is applicable to fuels with varying relative amounts of monocyclic and dicyclic components since the refractive index of the total aromatic fraction varies dramatically with the relative amounts of these two classes. High-performance liquid chromatography (HPLC) has been used routinely in this laboratoryG8 to separate jet fuels into three compound classes: saturates, monocyclic aromatics, and dicyclic aromatics. Quantitation of the separated fractions was initially accomplished by gravimetric analysis or gas chromatography. However, this was time consuming and the gravimetric analysis required complete sample recovery from the mobile phase. In an attempt to obtain quantitation at the time of separation with HPLC, the problem of determining an appropriate response factor for the aromatics is largely alleviated if the fuel is separated into the three compound classes. The refractive index of each aromatic fraction varies over a much more narrow range than that of the total aromatic fraction. We have employed a method that uses characteristic response factors for each class that are determined from “typical” fuels.9JO (2) Ashraf-Khrassani, M.; Levy, J. M.; Dolata, L. A. SFC Determination of Saturates and Aromatics in Petroleum Streams, Am. Lob. 1992, 24, 29. (3) Seng, G. T.; and Otterson, D. A. High Performance Liquid Chromatographic Hydrocarbon Analysis of Mid-Distillates Employing Fuel-Derived Fractions as Standards. NASA Tech. Memo 1983, No. 83072 (N83-19920). (4) Otterson, D. A.; Seng, G. T. GroupTypc Hydrocarbon Standards for HighPerformance Liquid Chromatographic Analysis of Mid-Distillate Fuels. NASA Tech. Pap. 1984, No. 2311 (N84-23774). (5) Miller,R.L.; Ettre, L.S.; Johansen,N.G.J. Chromozog. 1983,259,393412. (6) Solash, J.; Hazlctt, R. N.; Burnctt, J. C.; Beal, E.; Hall, J. M. In ACS Symposium Series 163; Stauffcr, Ed.; H. C., 1981; Chapter 16, pp 237-251. (7) Solash, J.; Hazlett, R. N.; Hall, J. M.; Nowack, C. J. Fuel 1978,57,521-525. (8) Hardy, D. R.; Hazlett, R. N. Analysis of Fuel Samples Provided by NAPC. Naval Research Laboratory Letter Report 6180-189:mld, March 22, 1983. (9) Sink, C. W.; Hardy, D. R.; Hazlett, R. N. Compound Class Quantitation of JP-5 Fuels by High Performance Liquid Chromatography/Diffcrential Refractive Index Detection, Naval Research Laboratory Memorandum Report 5407, September 13, 1984 (AD-A-145 754). (10) Sink, C. W.; Hardy, D. R.; Hazlctt, R. N. Quantitative Determination of Compound Classes in Jet Turbine Fuels by High Performance Liquid Chromatography/ Differential Refractive Index Detection, Part 2. Naval Research Laboratory Memorandum Report 5497, December 3 1, 1984. 0003-2700/94/036&1334$04.50/0
0 1994 American Chemical Society
1.65
Tablo 1. Moarurod Rofractivo 1nd.x.r of Compound C l a r u r from HPLC Rocovwod Fractkno of a Worldwld. Sunoy of Actual J.1 FwW
fuel 5-22 83-60 83-58 82-17 81-4 81-19 81-15 83-57 83-43 83-89 83-56 81-17 81-13 13-15 83-63 Pax-River 81-14 av
SD SD(%) range
RI of fractions monocyclic dicyclic saturates aromatics aromatics 1.4318 1.4363 1.4376 1.4376 1.4378 1.4382 1.4385 1.4387 1.4402 1.4405 1.4405 1.4405 1.4413 1.4414 1.4415 1.4424 1.4441 1.4393 k0.0028 k0.2 0.012
1.5111 1.5077 1.5128 1.5076 1.5074 1.5130 1.5074 1.5054 1.5135 1.5157 1.5157 1.5135 1.5066 NDb 1.5053 ND 1.5100 1.5102 k0.0036 A0.2 0.010
C C
C
1.6080 1.5950 C
1.5902 1.6065 1.6006 C C
1.6008 1.5902 ND 1.6000 ND 1.5902 1.5979 A0.0069 A0.4 0.018
A
n-Hexane
RI of whole fuel 1.4483 1.4499 1.4553 1.4572 1.4570 1.4554 1.4548 1.4502 1.4866 1.4515 1.4515 1.4730 1.4615 ND 1.4631 ND 1.4642 1.4586 A0.0102 A0.7 0.038
0 Ranked from loweat t o highest in term of saturates fraction RI. All RI data at 20-30 OC. Table is from ref 10. b ND, not determined. c Insufficient material available for measurement.
Numerous fuels were separated into the three classes, and the refractive indexes of the purified fractions were determined. The refractive index of the saturate fraction typically falls in the range of 1.43-1 -45. The monocyclic aromatics regularly have a refractive index near 1.51 while dicyclic aromatics are characteristically near 1.61, as shown in Table 1.lo Typical response factors for each class are determined, and quantitative analysis is obtained by using the peak areas of given volumes of standards with the appropriate refractive indexes. The rather large ARIs for the aromatic fractions, using pentane or hexane mobile phases, provide reasonably constant response factors for these classes. However, the small ARI and the large range of refractive indexes possible for the saturate class generate a much greater uncertainty in the quantitation of this class because the response factor of the calibration standard for this class must be matched much more closely to that of the unknown sample. Therefore, this approach always has a degree of uncertainty that cannot be eliminated. Since variations in refinery processes and feedstock sources will change the RI range of this class, the reliability of this method leaves much to be desired. There is undeniably a need for a quantitative method to determine hydrocarbon classes in all types of fuels that is accurate, requires no response factors, is relatively simple, and is applicable to both current and future fuels. This paper details a new method for obtaining compound class quantitation in jet fuels during HPLC separation without prior knowledge of the refractive indexes of these classes and thus requires no response factors. The method is applicable to all hydrocarbon fuels that can be separated into compound classes by HPLC. More importantly, it should also find application in general analysis schemes where the identity of the analyte may not be available prior to quantitation, such as in the application to environmental analysis.
I
1.35 -1
0
1
5 6 Areamnit Volume (millions)
2
3
4
7
8
Flguro 1. Data from Table 2 plottedto ghrearea/unt volume of sample vs analyte refractive Index In two different mobile phases. For hexane, the R I Intercept Is 1.3749,the measured value is 1.3756,the slope (k,) Is 3.198 X lod, and R 1s 0.999.For nonane, the R I Intercept Is 1.4038,the measured value Is 1.4038,the slope (k,) Is 3.348 X lod, and R2 Is 0.999.Note the nonparallel slopes. Slopes are used In eqs
5-9.
THEORY When one chromatographs a fixed volume of pure hydrocarbon samples, the signals produced from the response of a differential refractive index detector plotted vs the measured refractive index (Table 1) of these samples yields a nearly perfect linear correlation (Figure 1). (How to ensure this necessary linearity will be discussed in the Experimental Section.) This linearity is observed regardless of the mobile phase employed or whether the samples utilized belong to the same or different hydrocarbon classes. As expected, the intercept is the refractive index of the mobile phase employed because an analyte with the same refractive index as the mobile phase should give zero area or ARI. In addition, the slope is nearly independent of the choice of mobile phase. The small variation in slope that one does observe is most likely produced by the differences in back pressure caused by viscosity variations of the mobile phase. Dissimilar back pressures will produce a change in the pressure gradient in the flow cell. This can cause small distortions that result in slight differences in the signal for the same refractive index differential. It could also be because the two halves of the flow cell are at slightly different pressures, since the fluid in the reference side is static. This pressure differential may cause some distortion in the flow cell, which gives an apparent RI that is different from the value measured with a refractometer. The divergence between the slopes of this correlation for two mobile phases increases as the differences in back pressure increase and is almost zero for hexane and heptane, which produce nearly the same back pressure. For any given system/detector operating under identical experimental conditions, for any given mobile phase, the area is reproducible and the slope is readily incorporated in the calculations. Figure 1 shows a typical graph of the refractive index of analyte vs the area per unit volume from which the following equation is readily obtained: where nox is the refractive index of pure analyte x and ki is the slope of the line with mobile phase i. Aox,iis the area per Anal’lcal
Chemlstry, Vol. 86, No. 8, April 15, 1994
1335
unit volume measured for pure analyte x with mobile phase i and noi is the refractive index of pure mobile phase i. If a mixture such as dodecane, tert-butylbenzene, and l-methylnaphthalene is chromatographed, the area measured for each separate component x while mobile phase i is used is Aox,iand is equal to A , i = VxA0x,i where V, is the volume fraction of x in the sample. By combining eqs 1 and 2, the value of the refractive index of pure compound x is given by
(3) nox = kiAx,i/Vx+ noi The refractive index for each analyte can be expressed in terms of another mobile phase j as
nox = k,Axj/Vx+ noj
(4)
These two equations may then be solved for either nox or Vx to give
where Rx = kiAX,iIkJlXJ
(6)
Vx = (kiAx,i- k,Axj)/(noj- no()
(7)
and
In addition, because the sum of all the fractions is unity, a generalized form of eq 7 can be expressed as
which reduces to
Thus, it is possible that the volume fraction of a component in any mixture of organic compounds(or fuel) can be calculated from eq 9 solely from the peak areas measured for that component in the chromatograms obtained in two different mobile phases. Thequantitation can be accomplishedwithout knowledge of the refractive index of any analyte or even the mobile phases employed. The volume fraction of any analyte can also be calculated with eq 7, which uses the difference of the two areas measured for that analyte A , i and A,J, the refractive indexes of the mobile phases and the slopes ki and ki. One can also calculate the volume fraction of each class separately with eq 3 by substituting the values for A,J and the refractive index, nox,calculated for that class from eq 5 . The last two calculations depend only on the areas measured for that analyte. A final aid in verification is that the sum of the fractions will be unity or 100% for each sample, provided complete elution of all analytes is obtained. EXPERIMENTAL SECTION A modular HPLC system was used in this study. A typical setup consisted of the following or equivalent components. A 1338 Analytical Chemistry, Vol. 66, No. 8, April 15, 1994
Laboratory Data Control Model I11 dual piston pump was used to maintain the mobile phase flow rate at 3.00 mL/min. Separation of the fuel components was accomplished with a Whatman PXS 10125 Partisil PAC (chemically bonded alkyl amino cyano) analytical column (4.6 mm X 25 cm), which produced a back pressure of -800 psi with hexane and 1200 psi with nonane. The DRI detector was a Waters Model 401 that was maintained at 30 OC with a Fisher Scientific Model 800 Isotemp constant-temperature circulator operating at the factory preset setting of 30 OC, The injector was a Rheodyne Model 7410 equipped with a Model 7012 loop filler port, fitted with an internal 0.5-pL loop to maintain the reproducibility of the small sample size injected. This sample size prevented the flow cell from being overloaded for all but the most extreme samples (those with a saturate fraction of >85% and with an RI of 1.46) and allowed the detector to be operated at an attenuation of 4 X to ensure optimum linearity of the detector response over the entire electrical output range of -700 mV. A 500-pL syringe equipped with a Rheodyne Luerlock needle was used to fill the loop. With this configuration, the syringe needed to be filled only once for each sample and the loop was thoroughly flushed with 100 pL of sample to remove mobile phase, bubbles, and any residue from previous samples. The removal of all bubbles was particularly difficult if the syringe volume was 50 pL or less. Data collection and integration were accomplished with a PE Nelson 900 Series Interface equipped with the revision 5.1 software. The detector signal was also monitored with a H P Model 3390A integrator to obtain real-time hardcopy and to observe the base line between runs, since no signal can be observed between chromatograms with the PE Nelson equipment. Typical run time was 4 min, and data points were taken at a rate of 101s. All refractive indexes for the standards and fuel used in this report were obtained at -24 OC with an Abbe refractometer equipped with a digital readout for RI and temperature from American Optical. Mobile phases employedin this study included hexane,isooctane,and nonane, allof at least analytical grade. Since the refractive index of the mobile phase may vary slightly in different lots, one should ensure there is ample supply of a given lot for a given set of analyses. All fuels and standards were injected neat in all cases.
-
-
-
PROCEDURE Using the first mobile phase, chromatograms of standards and samples were obtained in triplicate and saved in a computer disk file. The mobile phase was changed and the refractive index of the second mobile phase exiting from the detector was monitored with the DRI detector until a stable base line was restored. When the refractive index of the mobile phrase exiting from the DRI detector was identical to the initial solvent RI, it was assumed that there would be no further elution of the first mobile phase from the column. This mobile-phase change required 1 h. Using the second mobile phase, the standards and fuel samples were re-chromatographed in triplicate and these data were also saved in a computer disk file. Since the order in which the two mobile phases are used is not important, the system could be left in this configuration for the next set of
-
~~~
Table 2. Measured Rdractlve Indexes d Standards VI tho Area per UnH Volume Used To Generate Flgure 1
a Y
4
k l NONAIIF
0 0
. .-
1
2
3
time (minutes)
Flgure2. Chromatograms of atypical fuel sample showing DRI signal. Peaks elute In order as saturates, monocyclic aromatics, and dicyclic aromatics. The higher DRI signal in hexane Is on the top and the lower DRI signal from nonane Is on the bottom.
APPLI CAT1ON Calculation of R, and noxfor a Compound Class. A series of four standard mixtures of dodecane, tert-butylbenzene, and 1-methylnaphthalene were prepared to test the applicability of the method over a wide range of composition. Two of the standards, S-555 and ,91055, wereunusually highin aromatic content and S-1021 and STD-X were prepared to represent the typical range of the dicyclic components normally found in jet fuels. The fuel samples were chosen to represent the range of saturate fraction refractive index typically found in current jet fuels as well as exhibiting a broad range of composition in both total aromatics and dicyclic aromatics. Thus, if the method is applicable to these samples, it is expected to be applicable to most fuels.
nox
hexane heptane isooctane nonane dodecane tert-butylbenzene l-methylnaphthalene
1.3756 1.3856 1.3899 1.4038 1.4200 1.4897 1.6137
Aoxj(hexane) 0.O00
3.284 X 4.441 X 9.070 X 1.488 X 3.617 X 7.449 X
106 l@ 106 108 106 106
Ao,j (nonane)
-8.447 X 106 -5.469 X 106 -4.051 X 106 O.OO0
5.317 X l@ 2.510 X 108 6.285 X 108
Table 3. Cornparkon of Refractive Indexes Calculated from Equation 5 and R, and Measured Values for Some Known8 and Jet Fuels’
4
samples. This approach maximizes the quantity of data that can be collected between changes of mobile phases. Typically, the same samples will give reproducible areas from one day to the next. However, we always verified linearity using standards each day that the method was run. Peak area measurements for standards may be streamlined by preparing two standards with different components for each class. Typically this consists of a mixture of equal volumes of a saturate and monocyclic aromatic and dicyclic aromatic components since this gives chromatographic peaks that stay on scale and these areas multiplied by 3 give the same area as when they are chromatographed separately. Integration precision of the chromatograms was greatly improved by careful choice of the starting and ending points of the individual peaks. The area of each compound class was determined at least twice or until the differences between successive measurements were less than 1% of the total area, except when the signal-to-noise ratio made this impractical. For most curves, this agreement was usually obtained in two measurements and quite often it was possible to get successive values that agreed to four or more significant places. The areas for each compound class from the three chromatograms was then averaged for use in any calculations. Figure 2 shows the overlay of the chromatograms of a typical fuel sample. This illustrates the decrease in signal observed when nonane is substituted for hexane as the mobile phase.
sample
sample
~OSAT
~OMCA
s-555(calc) S-1055(talc) s-1021(calc) STD-X (talc) av (calc) measd 5-22(talc) measd 82-17(talc) measd 81-14(talc) measd
1.4206 1.4208 1.4205 1.4205 1.4205 1.4200 1.4320 1.4318 1.4402 1.4376 1.4443 1.4441
1.4804 1.4839 1.4858 1.4822 1.4836 1.4897 1.5016 1.5111 1.5097 1.5076 1.5138 1.5100
~ODCA
1.5873 1.6052 1.6033 1.6013 1.5993 1.6137 1.4593b NAC 1.5974 1.6008 1.5863 1.5902
&AT
RMCA RDCA
2.682 2.661 2.712 2.692
1.368 1.352 1.340 1.360
1.154 1.140 1.141 1.143
1.971 1.279 1.493b 1.752 1.258 1.142 1.696 1.257 1.150
The typical values for R, for each fraction are calculated from eq 6. Measured RI values for hexane and nonane were used in the calculations. b This sample contained 1 % dicyclics and the area was too small to be measured reliably. NA, not available.
-
~
~~
~~~
The first step in the process was to plot the chromatogram peak areas (Table 2) produced by the standards A 0 , i (or A O , J ) vs the analyte refractive indexes nox,(and noxa Values of k,(or kj) and noi (or n o j ) were obtained from the slopes and intercepts of these lines, respectively (Figure l), from a least squares fit. The calculated refractive indexes of the mobile phases are identical to the measured values within the experimental limits as shown in Figure 1. Once the slopes and the intercepts (Figure 1) were determined, the compositional analysis was accomplished in the following manner. The value of R, for each class was determined by using eq 6. The values of R, are shown for the known mixtures and representative jet fuels in Table 3. In the saturate fraction, R, varied from 2.69 to 1.70 as the refractive index varied from 1.4200 to 1.4441. In the monocyclic aromatic fraction, R,, only changed from 1.36 to 1.26 for about the same ARI. The variation in R, for the dicyclic aromatic fraction was almost unmeasurable with approximately the same RI change. The R, value of each compound class was combined in eq 5 , with the calculated refractive indexes of the mobile phase, noi and no,, to calculate the refractive index of each compound class. These noxvalues were compared to the measured indexes for the standards and in this case, since the refractive indexes of the fuel fractions are known, to the corresponding values for these classes in “typical” jet f u e l ~ . Thus ~ J ~ one can check the validity of a set of data via the standards and spot fuels which may not be “typical”, especially in the saturate fraction. If the calculated and measured RIs of the knowns are in agreement, then the calculated values for the unknowns are also expected to be accurate (Table 3) and one can proceed to quantitation calculations. Analytical Chemistry, Vol. 66,No. 8, April 15, 1994
1337
tam 4.
Tho U u of Equatlon 0 To Calculatr V, or Vohuno Peront of Four Standard Mixtures and t h m R.pro$ontatlro Jot Fwb* vol %
SAT
MCA
DCA
33.3 32.4
33.3 33.0
33.3 34.6
50.0 49.9
25.0 25.2
25.0 24.9
76.9 78.3
15.4 14.7
7.7 7.0
82.0 81.7
16.5 16.6
1.5 1.7
78.0 76.9 78.4
21.0 20.7 21.6
1.0 2.4
21.0 21.0 32.8
8.0 7.9
(FIN
71.0 71.0 67.2
15.0 14.7 26.2
6.0 8.2
(FIN
79.0 77.1 73.8
sample
s-555 actual calcd S-1055 actual calcd 5-1021 actual calcd STD-X actual calcd 5-22 actualb calcd
(FIAI
82-17 actualb calcd 81-14 actualb calcd
0 The jet fuels were chosen from Table 1 to span the eatest range of saturates RI. The current standard method of F I f i s shown for comparison. These values were determined by HPLC using measured RIs of each fraction to determine accurate volume percent.
*
The most direct method for calculating the compound class composition is to use eq 9 to calculate V, in volume percent. This generates a normalized percent composition that depends only on the slopes, kt and kj, and the peak areas measured from the jet fuel chromatograms A , , and A,J. The method is straightforward and accurate, as shown in Table 4 for all samples. There are three additional alternative methods to obtain the composition from the data, which may be applied as a check of the results from eq 9. One method involves solving eqs 3 and 4 for V,. This employs the refractive indexes, nox, calculated with eq 5 (and 6), the appropriate peak areas Ax,i (or A,J), and the measured mobile phase refractive index noi (or noj). A second process involves using the calculated refractive indexes, nox, to determine values for Aox,,and Aoxjfor each compound class. Equation 2 and A , , (or A,$ can be used to calculate the volume fraction, V,, which is then converted to volume percent. These latter two methods give two determinations of each volume fraction, one for each mobile phase employed. If the calculated refractive indexes are accurate, one obtains the same results for both mobile phases. Equation 7 may also be used with the appropriate areas and mobile-phase refractive indexes to generate the volume fractions. Since this latter method involves data from both chromatograms, it represents an average of the values determined from eqs 2-4 employing data from both the hexane and nonane mobile-phase chromatograms.
1330 Analytical Chemlstty, Vol. 66,No. 8, April 15, 1994
These three methods yield “absolute”volume percents and may total more or less than loo%, depending on how well the refractive indexes are Calculated. If the calculated refractive indexes matched exactly the true refractive indexes of each class, the results obtained from the three alternate methods would total 100%and should be the same as those calculated with eq 9. If these “absolute”volume percents are normalized to loo%,the quantitation is equivalent to that calculated with eq 9 and the calculated composition is equal to the accepted values, within experimental limits, as demonstrated in Table 4.
The application of the technique to jet fuels tests the limits of both the range of composition and the ARI. The requirement of isocratic conditions limits the applicability somewhat. The task of finding two acceptable mobile phases may be facilitated by the use of blends of two or more components to achieve the necessary refractive index differences while maintaining appropriate chromatographic resolution to obtain the precision and accuracy required.
CONCLUSIONS The results of this study demonstrate the applicability of these concepts to the analysis of jet fuels, and the extension to other hydrocarbon fuels is believed to be straightforward. The method should have general application to other analyses where the identity of some components of a sample mixture are not available at the time of separation. The method is most accurate when the analyte refractive index is not greatly different from that of the mobile phases employed and when the composition of the components for analysis are all within 1 order of magnitude. This is illustrated in the dicyclics aromatic fraction in fuel 5-22, where it represented only about 1-2% of the fuel. If a value of 1.61 is used for the refractive index of this class and the composition is calculated by one of the alternate methods, the composition is 1%. Thisis in better agreement with thevaluedetermined p r e v i o ~ s l yThis . ~ ~ ~is ~probably due to the large uncertainty in measuring the areas of such small peaks. This uncertainty problem may be reduced by using larger sample sizes to more accurately determine peak areas or by the installation of a second injector with a loop size of 10 pL in series with the 0.5 p L loop. This would allow the analysis to be repeated immediately for small peaks and should enable more reliable determinations of R, to be made in such cases.
-
ACKNOWLEDGMENT C.W.S. thanks Michael Dawson and Blair Butler for their efforts as independent study students in helping to develop this technique. Received for revlew August 9, 1993. Accepted January 25, 1994.’
*Abstract published in Aduance ACS Absrracts, March 1,
1994.
