Expansion of dynamic range of flame atomic absorption spectrometry

Anal. Chem. 1009, 65, 1682-1888

1682

Expansion of Dynamic Range of Flame Atomic Absorption Spectrometry by an Efficient Flow Injection Dilution System Based on Dispersion of Microliter-Volume Samples Zhaolun Fang,+Bernhard Welz,' and Michael Sperling Department of Applied Research, Bodenseewerk Perkin-Elmer GmbH, W -7770 Uberlingen, Germany

An automatic on-line flow injection dilution system for flame atomic absorption spectrometry was developed based on the injection of low- and submicroliter samplevolumes and their dispersion in a mixing coil. Precise control of volumes was possible using computer-controlledstepper motordriven peristaltic pumps and small-bore Neoprene pump tubes. A more than 1000-folddilution with a precision of better than 2% (rad) (n = 15) was achieved using a sample volume of 0.7 MLand a mixing coil of 160-cm length and 1.3-mm i.d. Precision improved to 0.5-0.6% (rsd) when sampling times longer than 2 s were used. Sampling frequencies of 60-100/h, even for the highest dilutions, made the system highly compatiblewith routine application. A concentration of 7.2% Mg in an aluminum alloy was determined directly in the solution of 1 g of alloy in 100 mL. INTRODUCTION One of the drawbacks of flame atomic absorption spectrometry (FAAS)is its limited linear working range. Tedious multiple dilution steps therefore become an indispensable part of analytical procedures when analyte concentrations are high, and particularly when they differ within a wide range, This may degrade not only the efficiency but also the accuracy of an analysis, due to potential loss of the analyte, contamination, and introduction of operational errors during dilution. The integration of an efficient automated dilution system in FAA spectrometers to expand their dynamic range would undoubtedly enhance their performance. The combination of flow injection (FI) techniques with AAS has brought about a significant enhancement in the performance of the latter and has given rise to various new possibilities, including the introduction of samples with high dissolved solids content,' microsampling,2 control of interferences: on-line preconcentration and separation,4,6 and speciation.6 Many of these techniques showed considerable promise in routine applications.4Js8 Among the various

* To whom all correspondence should be addressed.

On leave from Flow Injection Analysie Research Center, Institute of Applied Ecology, Chinese Academy of Sciences, 110015Shenyang, China. (1) Fang, Z.; Welz, B.; Schlemmer, G. J . Anal. At. Spectrom. 1989,4, t

91. (2) Fang,Z.; Welz,B.;Sperling, M. J.Ana1. At. Spectrom. 1991,6,179. (3) Sperling, M.; Fang, Z.; Welz, B. Anal. Chem. 1991,63, 151. (4) Fang, Z. Flow Injection Separation and Preconcentration; VCH Verlagsgesellschaft: Weinheim, Germany, 1993. (5) Welz, B. Microchem. J . 1992,45, 163. (6) Sperling, M.; Xu, S.; Welz, B. Anal. Chem. 1992, 64, 3101. (7) -on, J. F. Spectrochim. Acta Reu. 1991, 14, 169. (8) Burguera,J. L., Ed. Flow Injection Atomic Spectroscopy; Marcel Dekker: New York, 1989. 0003-2700/93/0365-1682$04.00/0

techniques, FI on-line dilution for FAAS is one of the topics that has been studied earliest and reported most often. This is not surprising, since controllable dispersion, Le., dilution, is one of the cornerstones of FI analysis.9 The different approaches for dilution have been reviewed by Fanglo and, more recently, by Tyson.7 On-line dilution procedures based on dispersion of the injected sample in a mixing coil or mixing chamber have been proposed using merging streams,ll flow splitting,12 zone sampling,ls zone penetration," cascade dilution,ls and controlled dispersion.'s However, despite the progress achieved by several workers, and the relatively large number of different approaches available, hitherto the techniques did not appear to be efficient, reliable, precise, and versatile enough to satisfy the requirements of a routinely applicable procedure when large dilution ranges were required. In this study an attempt was made to develop an efficient and reliable, yet simple and versatile wide-range on-line dilution system for routine FAAS based on the introduction of sample microvolumes using commercially available equipment.

THEORETICAL CONSIDERATIONS In early studies9 on the effect of variations in the injected sample volume on dispersion, an exponential relationship was observed between the dispersion coefficient D and the sample volume V, according to

D = 1- exp(-Vk)-' (1) where k is a constant determined by the experimental conditions. The dispersion coefficient D is defined as cdc, where c, is the original concentration of the sample before dispersion and c the concentration of that fluid element of the dispersed sample zone where the readout is taken. D is therefore a value which is greater than unity and is equivalent to the dilution factor. It has been indicated that the rising edges of all curves described by eq 1coincide, irrespective of the sample volume injected. Within the first portion of the rising part correspondingto less than 50% of cotor a dispersion coefficient larger than 2, an almost linear relationship could be obtained between V and c . ~Therefore, it should in principle be possible to obtain large dilution factors by decreasing the volume of the injected sample. In fact, this was the first rule for dispersion control proposed by Ruzicka (9) Ruzicka, J.; Hansen, E. H. Flow Injection Analysis, 2nd ed.; John Wiley: New York, 1988. (10) Fang, Z. In Flow Injection Atomic Spectroscopy; Burguera,J. L., Ed.; Marcel Dekker: New York, 1989; p 103. (11) Zagatto, E. A. G.; Krug, F. J.; Bergamin, F. H.; Jorgensen, S. S.; Reis, B. F. Anal. Chim. Acta 1979, 104, 219. (12) Mindel, B. D.; Karlberg, B. Lab. Pract. 1981, 30, 719. (13) Reis, R. F.; Jacintho, A. 0.; Mortatti, J.; Krug, F. J.; Zagatto, E. A. G.; Bergamin, F. H.; Pessenda, L. C. R. Anal. Chim. Acta 1981,123, no, LLI.

(14) Zagatto, E. A. G.; Gine, M. F.; Fernandes, E. A. N.; Reis, B. F.; Krug, F. J. Anal. Chim. Acta 1985, 173, 289. (15) Whitman, D. A.; Christian, G. D. Talanta 1989,36,205. (16) Sherwood, R. A.; Rocks, B. F.; Riley, C. Analyst 1985,110,493.

@ 1993 American Chemlcal Society

ANALYTICAL CHEMISTRY, VOL. 65, NO. 13, JULY 1, 1993

a

and Hansen in their pioneering studies on the basic principles of FI.17 As could be seen from eq 1, another means for increasing the dispersion would be to decrease the k constant of the dilution system. This might be achieved by using mixing coils with larger tube diameters and lengths. In practice, however, FI dilutions exclusively based on sample dispersion were limited to relatively low dilution factors of not more than 10. When Vis expressed in the form of nVIp, where Vlp is the sample volume producing a dispersion coefficient of 2, and n the number of V1,is comprising V, eq 1may be transformed into 1/D = 1- 2"

f = 6OP/2VD = 3OP/VD

V

-

C

I

P1

P2 ON

ON

L-(2)

It could be derived from this equation that for a D (or dilution factor) of 1000, n = 0.001 44, implying a sample volume of less than 1.5 pL for a system where D = 2 could be obtained by injecting a 1000-pL sample. Even for a target dilution factor of 100, where n = 0.0144, the injected sample volume should be less than 15 pL, which is below the minimum normally manageable by injection valves equipped with sample loops. Admittedly, V1p could be increased by increasing the length and capacity of the mixing coil, i.e., improving the dilution proficiency of the system, however, only at the expense of a decreased sampling frequency. If a 30-pL sample, usually the smallest sample volume introduced by an injection valve, were diluted 1000-fold, the resulting diluted sample bolus would have a volume of 30 pL, assuming homogeneous distribution of the analyte over the length of the bolus. The volume would be at least twice as large if the skewed-Gaussian distribution, normally encountered for FI peaks, were taken into consideration. A rough estimation of the possibly highest sampling frequency f (l/h) for a dilution system with a moderately skewed peak output signal could be made using eq 3. (3)

with V expressed in milliliters and P, the carrier flow rate, in milliliters per minute. For an injection volume of 30 pL, a carrier flow rate of 4 mL/min, and a target of 1000-fold dilution, the highest achievable sample throughput would be about 4/h, which obviously is not acceptable for routine analyses. The sampling frequency would be decreased even more if dilution chambers were used that produce extremely skewed output signals. Equations 1-3 suggest that, with a simple dispersion system built from mixing coils, high dilution factors and sampling frequencies could be achieved only if small sample volumes in the low- or submicroliter range were introduced. The dispersion of the system should also be large enoughto produce a dilution factor of D = 2 with a 0.5-1-mL sample injection. The dilution system developed in this study is based on these principles. EXPERIMENTAL SECTION Instrumentation. A Perkin-Elmer Model 4100 atomic absorption spectrometer was used without background correction in the flame flow injection mode with a flow spoiler in the spray chamber. A magnesium hollow cathode lamp, operated at 6 mA, was used and the wavelength set to 285.2 nm. The acetylene and air flow rates were set at 1.9 and 8.0 L/min, respectively. The nebulizer uptake rate was adjusted to 6 mL/min, the lowest rate that gave optimum response in the conventional mode of sample introduction. (17) Ruzicka, J.; Hansen, E. H. Flow Injection Analysis, let ed.;John Wiley: New York, 1981.

1689

b

INJECl

-

-

*W

L

C

-

---

w

~

I

The time-resolved absorbance signals were displayed on the high-resolution graphic screen and were printed by an Epson Model EX-800 printer. A time constant of 0.7 s was used for peak height absorbance (A&, and an integration (read) time of 30-50 s, with a read delay of 10-15 s, depending on the duration of the peak, was used to evaluate integrated absorbance (Aht). A Perkin-Elmer Model FIAS-200FI system with arotary injector valve with four channels at the stator and five at the rotor was used. The FI manifold is shown schematically in Figure 1. In some studies, the sample inlet channel on the rotor of the valve preceding the sample-loopchannelon the stator was made thinner by threading into the 1-mm-diameter channel a piece of 0.5mm4.d. 1.6-mm-0.d. Micro-Linetubing (Thermoplastics,Sterling, NY) which was stretchedthin. After the threaded tube was pulled to produce a tight contact between channel and tube, the ends of the tube were cut flush to the surface of the valve rotor. This resulted in a channel diameter of -0.3 mm which minimized free migration of sample into the sample loop during actuation of the valve (see Results and Discussion). The sample loop was made from a 12-cmlength of I-mm4.d. poly(tetrafluoroethy1ene) (PTFE) tubing with flanged ends and connected to the valve stator by threaded fittings. The volume of the loop, including the valve channel,was 100pL. For the sampleuptake capillary, a 10-cm length of 0.35-mm-i.d. PTFE tubing was connected to the valve rotor. Pump 1of the two programmablestepper motordriven peristalticpumps, used for samplemetering, was equipped with Ismaprenepump tubes (Ismatec,Wertheim, Germany)with a 0.25-mm i.d. and 2.07-mm o.d., while pump 2 was equipped with ordinary Tygonpumptubes. All pump tubes were lubricated with a small amount of silicon oil. The mixing coils were made from 1.3-mm4.d. Micro-Line tubing, coiled with an outer diameter of 10 cm and mechanically fixed on the tray of the FI system to minimize movement of the coil during turning of the valve rotor. The coils were connected to the valve rotor by a 10-cm length of 0.6-mm4.d. Micro-Line tubing and to the nebulizer by 100 cm of the same tubing. The latter was either coiled to produce a 3-cm-diameter coil or knotted into a three-dimensionally disoriented configuration in order to improve mixing conditions.2 The thin conduit also acted as a flow restrictor to prevent formation of air bubbles in the mixing coil or chamber, which might have degraded the precision of measurement.

-

1684


Table I. Sequence of Operation for the FI Dilution System. pump speed time (rpm) flow rate valve step function (a) P1 P2 (mL/min) position 1 2 3 4

valveturn 1 0 sampling 1-10 10-100 baseline offset 2 0 correction sampleto 40 0 detector sample flush

80 80 80

5.7 5.7 5.7

fii

80

5.7

injectb

fill inject

0.24

0 Pump tubes were 'blue-yellow" for carrier and "blue-orange"for sampling and sample flush. b Read cycle initiated with 10-8 delay.

A variable-volume mixing chamber, similar to that described by Beinrohr et al.t8was used in some of the dilution studies. It was made from a 20-mL plastic disposable syringe, and a homemade 12-mm-long,1.5-mm-0.d. stir bar was inserted inside the chamber to improve mixing. The mixing chamber was fiied in the center of, and 3 cm above, the plate of a magnetic stirrer. The mixing chamber was connected to the valve by 30 cm of 0.3-mm-i.d. Tygon tubing and to the nebulizer using the same flow restrictor as described above. Reagents and Standard Solutions. Demineralized water was used as carrier for all studies unless specified otherwise. Degassing of the water was not necessarywhen the flow restrictor described above was implemented. Standard solutionsfor magnesium were obtained by diluting a 1OOO mg/L stock solution, prepared from Titrisol standard concentrates (Merck, Darmstadt, Germany), with water. For dilution studies of highly concentrated solutions, the 1OOO mg/L stock solution was used directly. For the study of matrix effects, a 1OOO mg/L solution of MgC12.6Hz0 in water was used. A solution of an aluminum alloy standard reference material, BAM-206 (Bundaaanstaltfih MatarialprOfung,Berlin, Germany), was made by dissolving 1.OOO g of metal in 10 mL of 6 mol/L hydrochloric acid and dilutingto 100mL with water. The sample was analyzed using simple aqueous standard solutions but with a 0.5% (w/v) La carrier, which was prepared by dissolving 5.864 g of lanthanum oxide in 250 mL of hydrochloric acid and diluting to 1 L with water. Procedures. The sequenceof operation of the dilution system is shown in Table I. During the warmup period, the sampling pump (pump 1)was run at increased speed, with the valve in "fill" position, aspiratingwater into the sample loop through the sampling capillary and fiiing the entire pump tube with water. Care should be taken not to tighten the pressure adjusters for the pump tubes more than necessary. Under normal conditions, pump 1was stopped for more than 90% of the operation time. Under such conditions, excessive pressure on the pump might seriously deform the tubing, degrading the performance of the dilution. During the 10-8prefill period, which was only actuated when a new sample was introduced, the valve was in the 'inject" position. The sample takeup capillary was flushed with the sample to be analyzed, which was appreciated by pump 2, displacing the water or the previous sample from the tube. In sequence 1, which lasted only for 1s, the valve was turned to the fill position. In sequence 2, pump 1was actuated at a predetermined rotation rate for a defiied period of 1-10 8, during which the sample in the takeup capillary and connecting valve channel was metered into the sample loop. Pump 1was stopped before the valve turned to the inject position in sequence 3, when transport of the metered sample to the mixing and nebulization systems was initiated by the carrier stream propelled by pump 2. This sequencelasted only 2 sand was introduced only because the sequence immediately preceding the read cycle of the measurement should not be shorter than 2 s in order to define the baseline level before the reading was initiated, and sequence 2 might last only 1 s when the highest dilution factors were

required. In sequence4,which was the main injection sequence, valve and pump conditions were exactly the same as for (3).The injected sample was dispersed in the mixing coil or chamber,and the diluted sample was introduced into the nebulizer-burner system. The read cycle was initiated at the-beginning of this sequence with an appropriate time delay, and the peak height and integrated absorbance were evaluated. Simultaneously,the sample was taken up through the sample capillary by pump 2 to ensure the identity of the sample composition at the interface of the rotor and stator of the valve for the next injection. When a different sample was introduced for the next injection, the sample might be changed at the beginning of the last sequence, in which case the prefii time for the ensuing samples may be shortened or totally deleted, depending on the length of the injection sequence and the flow rate of the sample-flush pump channel. The system was calibrated injecting identical volumes of concentrated standard solutions.

RESULTS AND DISCUSSION Practical Considerations in the Design of the Dilution System. As indicated in the theoretical section, the high dilution factors pursued in this study were based on the introduction of sample volumes in the low-microliter to submicroliter range. Apart from special hardware requirementa for making this possible, the feasibility of such an approach relied strongly on the reproducibility of the sampling process. The conventional approach of sample introduction by first filling the sample into a loop of defined volume and then injecting the sample by a carrier stream, following the actuation of an injector valve, was not suitable for that volume range. Although injector valves with fixed sample bores of a few microlitershave been described, such an approach would be associated with losses in versatility, and variation of the dilution factor would not be convenient. The controlled dispersion system developed by Sherwood et ala,'@which did not use an injection valve and which was designed for microvolumes of clinical samples, appeared to be capable of providing a solution to this task. The system involved the use of a stepper motor-driven peristaltic pump and a movable sample probe, all under computer control. After an equilibration period, the pump was stopped and the sample probe immersed into the sample solution. The pump was restarted, moving through a predetermined angle of rotation, drawing a small volume of sample into the probe, and stopped again,and the probe was moved into the carrier solution. After the pump was restarted, the metered sample was transported to the detector by the carrier stream. The most significant advantage of the controlled dispersion system was in the opinion of the inventor the ease with which the sample volume could be varied.19 However, an important feature of this system was the use of a single pump channel for sample loading and carrier flow. This feature would not pose any problems as long as the sample volumes were relatively large. However, it appeared questionable that any single pump channel could provide a flow rate of 4-8 mL/min (the normal carrier flow rates for FI-FAAS) and at the same time meter samples a t the submicroliter or low-microliter level with a reproducibility of 1-2% rsd (the precision expected for a routine dilution system) even when a variablespeed pump was used. With the 0.74-mm4.d. pump tube used in the controlled dispersion system, the linear distance of roller movement is less than 6 mm for a 2.5-pL sample for which a precision of 1.9% red was reported. Although the system was capable of introducing samples as small as 1.25 pL, no precision data were given for that low level. Another potential danger in using a single pump line for sample and ~

(18) Beinrohr, E.; CsBmi, P . ; W n ,J. F. J. Anal. At. Spectrom. 1991, 6, 307.

(19) Sherwood, R. A.; Rocks, B. F. In Flow Injection Atomic Spectroscopy; Burguera, J. L., Marcel Dekker: New York, 1989; p 259.


carrier was the occasional entrainment of a small air segment during the movement of the probe between the sample and carrier containers, which obviously degraded precision. Although the system might be sufficiently reliable for dilution factors of -100, it was considered not robust and reliable enough for the realization of dilution factors of -1000. Nevertheless, the use of a stepper motor-driven pump for the metering of small sample volumes was a feature that proved successful in the controlled dispersion system, and the idea was adopted in a modified form in the system developed in this work. Recently, the concept of sequential injection wm incorporated into the traditional injection modes of FIA and considered to be one of the central elementa in the second coming of F1.m The carrier, sample, and reagent solutions were sequentially metered into a holding coil and injected into the mixing system by flow reversal. Although a valve was still required in such a system, this approach obviated the need of a sample loop and introduced more flexibility in the control of reagent and samplevolumes. Thisfeature might be used to advantage for the design of a versatile broad-range dilution system. The system in this work was developed using commercially available equipment, adapting some of the favorablefeatures of the previously mentioned systems. This involved many practical considerations, such as the design of the sample metering system, technical considerations associated with sample changeover, and optimization of dimensions and configuration of the mixing system. The following specific features were incorporated into the design of the system to fulfill the aim: (i) In order to make possible the metering of low- and submicroliter samples using conventional injector valves without sacrifice in flexibility, the dilution manifold was designed in such a way that the sample was metered into a conventional sample loop of 100 pL, but without the need to fill the loop or flush it with the sample before filling. The sample volume introduced into the loop was in the range of 0.7-35 pL, so that the loop was filled with the sample to only one-third at most. (ii) A separate pump tube each was used for the carrier stream, sample metering, and sample flushing so that the tube dimensions and flow rates could be optimized independently without compromise and sample change be achieved without fiiing or flushing of the sample loop. The sample flush pump channel was reserved for flushing of the sample takeup capillary only. (iii) In order to ensure better precision in the metering of sample microvolumes, Neoprene pump tubes with better elasticity than those made of Tygon, and with the thinnest available bore diameter of 0.25 mm, were used. The elasticity and large wall/bore ratio (7.3) minimized deformation of the pump tube even after they were pressed for extended periods with the pump stopped. The small bore size also made possible meteringof sample microvolumeswith larger rotation angles of the pump, which improved the reproducibility. (iv) A computer-controlled variable-speed stepper motordriven peristaltic pump was used to ensure precise movement of the rollers during the metering of samples. A single rotation of the pump was composed of 200 steps, corresponding to a rotation angle of 1.8' for each step, and the go-atop timing of the pump was precise to the order of milliseconds. (v) In order to guarantee the accuracy of the set sampling time, the sampling period was initiated only after the valve turn had been completed. For this reason, a separate sequence of 1-8 duration was included in the program, exclusively for changingof the valve position without disturbing the sampling

-

(20) Ruzicka, J. Anal. Chirn. Acta 1991,261,l.

1885

I n0. 0I "

I

10

20

30

40

50

60

70

80

Time (SI

Flgure 2. Recordings of dilution peaks for 1000 mg/L Mg obtained with 0.7-pL sample and 4.3 mL/min carrier flow: (a) mixing coil 120 cm long, 1.3-mm 1.d. (1.8 mL); (b) 1.5-mL mixing chamber.

process (the sample metering pump remained stopped during this sequence). (vi) The diameter of the valve channel connected to the sample uptake capillary was decreased as described in the Experimental Section to prevent migration of sample into the sample loop during turning of the valve, particularly when the sampling matrix was very much different from that of the carrier. This produced a particular interference effect which is discussed under Matrix Effecta. (vii) After having compared the overall performance of a mixing coil and a mixing chamber, the former was preferred for ita simpler and more robust construction and its higher efficiency. Details of the comparison are given in the section discussing the design of the mixing system. (viii) A knotted reactor made of thin-bored tubing positioned between the mixing coil and the nebulizer was used to upgrade the precision as described in the Experimental Section. Optimization of Experimental Parameters for the Dilution System-Design of the Mixing System. Mixing systems intended for dilution were evaluated not only by the dilution factor achievable but also by their sampling frequencies and short- and long-term precision. A comparison of the signals generated by a mixing coil and avariable-volume mixing chamber resembling that proposed by Beinrohr et al.la is shown in Figure 2. Both mixing systems had approximately the same dead volume (1.5 mL), identical sample volumes, and flow parameters. The dilution factor for the mixing chamber was higher by a factor of 2.8 compared with that of the mixing coil, but the duration of the peak (baseline to baseline) was a factor of 4 longer. The sample volumes used in this study (0.7 pL) were much lower than those used by Beinrohr et al. (15 pL/min), and therefore, the same dilution factors were achieved with a lower mixing chamber volume and the measuring time was much shorter (-2 min for a 3000-fold dilution). Nevertheless, in general, our findingswere in accordancewith theirs in that the working range could be enhanced by 2-3 orders of magnitude with mixing chambers, but also that prolonged measuring times of up to 10min were required for high dilution factors. Despite the substantially shorter measuring time obtained in this study, it was not considered worthwhile to pursue the higher dilution factor at the expense of the significantloss in sampling frequency. An additional drawback of the mixing chamber approach was that air bubbles were produced in the mixing chamber upon intensive stirring, when the carrier solution was used without degassing, even when a flow restrictor was added downstream. This increased the noise level and degraded precision. Generation of air bubbles could of course be avoided by degassing of carrier solutions, which would have added, however, to the complexity of the procedure. The use of mixing chambers was therefore abandoned. Mixing Coil Length. In order to increase sample dispersion, tubing with a relatively large inner diameter of 1.3 mm was used for the mixing coil. The diameter of the coil


1888 0.7

200

1a

0.5 \ C

150 Y-

0.4

r c

s

Y

z

8

100

6 0.3

m .e

50

Time ( s )

0

L

50

Flgure 9. Influence of mixing coil length (1.3-mm Ld., l k m coil dlameter)on signals for 100 mg/L Mg standard solution using 1.4-pL Injectlons (2 s, 10 rpm) and a carrler flow rate of 5.7 mL/mln: (a) 0-, (b) 30-,(c) 60-, (d) 90-, (e) 120-, (t) 170-, and (9) 2 1 k m coil length. 900 1 f l 900

0

40

I

I

I

60

70

80

Pump

rl’/ 0

0

I

50

rotation speed

I

I

3.0

4.0

tr.p.m.) I

5.0

I

6.0

Carrier f l o w rate (mL/min) Flgure 5. Influence of carrier flow rate on peak height absorbance, Awl and on sampling frequency, f: 100 mg/L Mg standard solutlon, 5.9-pL sample volume, coil length 120 cm.

Table 11. Effects of Sampling Pump Rotation Speed on D and Precieion (rsd). PUP

10 25 40 70 100

, 0

50

150 Mixlng c o i l length tcm) 100

‘ 0

200

Flgure 4. Influence of mlxlng coil length on dispersion factor, D, and sampllng frequency, f. For experimental parameters see caption of

Figure 3. was also relatively large in order to decrease secondary flows. This should result in decreased radial and enhanced axial dispersion of the injected sample. Seven different coil lengths were investigated, and the signals obtained for 1.4 pL of a 100 mg/L Mg standard are shown in Figure 3. As expected,higher dilution factors were obtained for the longer coils. The influence of the mixing coil length on D and f is shown in Figure 4. It was interesting to observe that a 100-folddilution might be obtained even without a mixing coil. This dispersion effect was due to contributions from the sample loop, the valve channels, and the connection lines when the small sample volumes used in this study were injected. Unexpectedly, however, an almost linear relationship was obtained between the coillength and the dispersion coefficientD, which was not consistent with the observations of Ruzicka and HansensJ7 that D varies linearly with the square root of the coil length. This appeared to be a phenomenon associated with the extremely small sample volumes and large reactor volumes used in this work and which deservesfurther studies. Figure 4 also shows that the samplingfrequenciesdecreased with an increase in mixing coil length however, even with a 210-cm coil, which produced an -9OO-fold dilution with a 1.4-pLsample, a theoreticalf of 80 might be reached. Higher f values could be achieved with some sacrifice in dilution factors by using shorter coils. With the consideration of maintaining a sampling frequency of 100/h at a carrier flow rate of 5.7 mL/min, the coil length adopted in this study was typically 170 cm. Carrier Flow Rate. The effects of the carrier flow rate on D and f were studied using a 120-cm length of coil and 4.9

sample 0.7 1.8 2.8 4.9 7.0

0.045 0.122 0.211 0.323 0.400

1.74(n= 7) 1.81 (n = 10) 0.74(n = IO) 1.04 (n = 8) 1.70 (n= 9)

0.430 1.21 2.10 3.24 4.05

1.83 (n = 7) 1.30(n = 10) 0.80(n = 10) 0.79(n = 8) 1.40 (n = 9)

1330 492 284 185 150

0.75 2.0 3.5 5.4 6.6

a Sample, 100 mg/L Mg; eampling time, 1 e; coil length, 160 cm, 1.3-mmi.d.; carrier flow rate, 5.7 mL/min; samplingtube, 0.25-mmi.d. Ismaprene.

pL of sample. The resulta that are presented in Figure 5 show very little influence from the carrier flow rate on D above a flow rate of 3.6 mL/min. The sampling frequency, f, estimated from the baseline-to-baseline time of the peaks under different carrier flow rates, was affected much more. However, theoretical f values of 90-100/h were achievable with a flow rate of -4 mL/min; the choice of flow rate was therefore not critical, but analytical results might be less affected by flow rate changes when carrier flow rates were higher than 4 mL/min. SampleMeteringTimeand Pump Speed. In the present system,the samplevolume might be changed by varying either the metering time, t, or the speed, t , of the sampling pump. Table I1 shows the dilution factors and precision for Apwlr andAbt evaluations obtained from varying r using the shortest sampling time of 1 s. A roughly linear relationship was observed between r and 1/D, and precision was best for medium rotation speeds. A similar relationship was obtained between the sampling time t and 1/D,as can be seen in Table 111. Precision was very good, particularly in integrated absorbance, for sampling times longer than 2 s. However, at the lowest pump speed of 10 rpm, precision degraded significantly for short sampling periods, particularly for l-s sampling. This is quite understandable, since with this set of conditions the pump only rotated through a 60° angle, within which the position of the pump rollers could affect the delivered volume. The precision might be improved by controlling the roller position at the beginning of each sampling cycle, as proposed by Sherwood et al.,le but this was not possible with the software used in this study. The effects from simultaneously varying t and r on D and on the precision of A m measurements are shown in Table


Table 111. Effects of Sampling Time on D and Precision (rad)* rsd rsd sampling sample time vol 1/D (n = 8) Aet (n = 8) (%) (8) (%) D (XlW (8) (PL) A+ 1.0 2.0 3.0 5.0 7.0 10.0

0.7 1.4 2.1 3.5 4.9 7.0

0.064 0.134 0.204 0.340 0.459 0.625

2.2 1.20 0.82

0.90 0.73 0.91

0.53 1.34 2.06 3.45 4.77 6.66

2.1 1.05 0.53 0.51 0.53 0.54

938 448 294 176 131 96

1.1 2.2 3.4 5.6 7.6 10.4

Sample, 100 mg/L Mg; pump speed, 10 rpm; coil length, 150 cm, 1.3-mm i.d.; carrier flow rate, 4.3 mL/min; sampling tube, 0.25-mmi.d. Ismaprene.

1687

Table V. Linear Working Range and Dilution Factor (0) for Different Sampling Parameters. linear dilutn working samplingparams ,,,bration t r V curveslope corr factor, range (8) (rpm) (rL) (L/mg) coeff D (mg/LMg) conventional 0.60 0.9999 1 0.005-0.5 10 2 1

100

50 10

70 7 0.7

0.044 0.0051 0.00040

0.9999 0.9999 0.9998

13.6 117 1500

0.05-5 0.5-50 10-500

Coil length, 170 cm, 1.3-mm i.d.; carrier stream, 5.7 mL/min; sampling pump tube, 0.25-mm4.d. Ismaprene. 0.4

a

1

Table IV. Dispersion Coefficients D (Dilution Factors) Obtained under Various Combinations of Sampling Time (t) and Pump Rotation Speed (r)for the Same Sample Volume (- 7 pL)* t (8) r (rpm) A d D rsd (%) n 1.0 2.0 4.0 10.0

100 50 25 10

0.535 0.651 0.665 0.659

112 92 90 91

1.02 0.85 0.95 0.91

16 14 13 12

Sample, 100 mg/L Mg; coil length, 120 cm, 1.3-mm i.d.; carrier stream, 4.3 mL/min.

0 0

IV. Owing to the relationships deduced from Tables I1 and 111, the dilution factor (or D) might be fairly well predicted for different combinations of t and r, once D is known for a single set o f t and r from calibration. This was verified by the results in Table IV, where different combinations of r and t (rt constant) were used to produce approximately the same dilution factor. Matrix Effects. It was expected that as long as matrix effects do not occur under conventional analytical conditions with manual dilution, interferences should not be encountered with the FI dilution system. The only factor that might have an influence on sample dispersion is a different viscosity of sample and standard solutions. However, it was shown by Tyson et al.21 that, at least in FAAS applications, such effects were usually negligible. Nevertheless, early in our studies we observed a strong enhancement effect of 100% in A@ and Aht for magnesium in 1mol/L nitric acid solution compared to aqueous standards if 0.7-pL samples were injected. The effect decreased with increasing sample volume, and only 10 and 5 % enhancement was observed in A w and Abt, respectively, for 7-pL samples. This phenomenon disappeared, even for the 0.7-pL sample, when a carrier solution of the same acidity as the sample was used, suggesting that the effect might be associated with diffusion phenomena. This was verified by investigating magnesium standards in 1 mol/L "03 with water as the carrier, maintaining the sampling time of 1-10 s, but setting the pump rotation speed to zero. Surprisingly, a reproducible peak was obtained even with a 1-s sampling time, with an A w and Aht equivalent to a 0.7-pL injection of an aqueous magnesium standard with the same concentration. The peak increased with increasing sampling time, but not proportionally, and a 70% higher signal was obtained with a 10-fold increase in sampling time. On the other hand, when the aqueous standard was introduced under the same conditions, the resulting peak was negligible. The effect was therefore considered to be mainly related to an entrainment of the sample from the valve channel connected with the sample takeup capillary into the sample

-

(21) w o n , J. F. Analyst 1985, 110, 481.

0.5

1

Time (SI

50

b

0 0 Time t s ) 50 Flgure 6. Superimposeddilution peaks using 120-cm mixlng coil and 4.3 mL/min carrier flow rate: (a) 4.2 pL (3 s, 20 rpm) of 100 mg/L Mg; 15 signals superimposed; (b) 0.7 pL (1 s, 10 rpm) of 1000 mg/L Mg; 10 signals superimposed.

loop at the interface of two valve channels during the turning of the valve from injection to fill position. The effect was significantly enhanced by large channel bores in the injection valve and a large difference in matrix composition, which resulted in an increased diffusion speed. The standard injector valve used in the first stage of the study was therefore modified as described in the Experimental Section to decrease the diffusion area. This measure proved to be quite effective in eliminating the effect with a decrease of the spurious signal by a factor of 20. The amount of sample entrained at an interface of water and 1 mol/L HN03 was estimated to be only -0.03 pL with the modified valve, and an enhancement of less than 5 7% might be expected with the acid sample even with simple aqueous standards and 1-s sampling. Performance of the Dilution System. The linear regression equations of calibration curves obtained under three different sets of sampling parameters are shown in Table V, together with the equation for conventional calibration. The slope ratios imply lo-, 100-,and 1000-fold dilutions of the original sample. The reproducibility of the dilutions under the conditions 1s, 10 rpm and 3 s,20rpm is illustrated by 10and 15overlying

1888


FI peaks, respectively, in Figure 6a,b. The peaks were randomly taken from a set of 15and 30 injections made within 20 and 30 min, respectively. The precision of the measurementa was 1.8 (Ape& and 1.5% (Abt) rsd (n = 15) and 0.90 ( A d and0.54% (Aht) rsd (n= 29,withoneresultdiscarded), respectively. Generally, better precision was obtained using peak area evaluation. The dilution system was further tested by analyzing a standard aluminum alloy (BAM-206) with a certified magnesium content of 7.17 % using the 170-cm mixing coil. One gram of sample was dissolved in 100 mL of acid as described in the Experimental Section and analyzed without further dilution using 2 s, 10 rpm (1.4 rL) sampling. The magnesium content determined was 7.20 f 0.12% (n = 6) in the metal (720 mg/L in solution), which is in good agreement with the certified value. Although a sampling frequency of 60lh was used in most of the studies in order to collect and print the required data between samples, a frequency of 100/h was readily achieved in later studies.

CONCLUSIONS High dilution factors of 2-3 orders of magnitude for FAAS were achievable with good precision and high sampling frequencies by a FI dilution system with a mixing coil through the introduction of low- and submicroliter volumes of sample using computer-controlled stepper motor-driven peristaltic pumps equipped with small-bore Neoprene pump tubes. Owing to the ease and convenience with which dilution factors could be varied within large concentration ranges, the system might be readily developed into an intelligent dilution system with which the dilution factors were optimized and controlled by the computer. The almost linear relationship between the dilution factor and sampling timelrotation speed might as well form the basis for an intelligent calibration system. Further work will be conducted along such lines.

RECE~VED for review December 16, 1992. Accepted March 22, 1993.

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