producing a stable dehydrated compound (Fig. 3, Scheme III). A very weak peak for the loss of only H20 is observed in the mass spectral data (Table I). Part of the intensity of this peak could be contributed from H20 losses from other portions of the maduramicin a molecule. The fur- ther abundant loss of CO2 from the stable f~-olefin or ~/-olefin carboxylic acid group is unlikely in the FAB experiment, as evidenced, for example, by the weak ion in lasalocid, corresponding to the loss of C02 from a benzoic acid moiety. ~ Therefore, abundant consecutive losses of H20 followed by CO2 (Fig. 3, Scheme III) are very unlikely.
ACKNOWLEDGMENT
The authors greatly appreciate the technical assistance of Robert Sheridan in providing the computer resources for this work as well as instructive comments on the manuscript.
1. M. M. Siegel, W. J. McGahren, K. B. Tomer, and T. T. Chang, Biomed. Envir. Mass Spectrom. 14, 29 (1987).
2. W.J. Hehre, L. Radom, P. v. R. Schleyer, and J. A. Pople, Ab Initio Molecular Orbital Theory (John Wiley, New York, 1986).
3. J. Clark, A Handbook of Computational Chemistry (John Wiley, New York, 1985).
4. J. S. Binkley, M. J. Frisch, D. J. DeFrees, K. Raghavachari, R. A. Whiteside, H. B. Schelgel, E. M. Fluder, and J. A. Pople, GAUSSIAN82 Revision H Version (Carnegie-Mellon University, Pittsburgh, 1985).
5. C. M. Liu, T. E. Hermann, A. Downey, B. La T. Proser, E. Scheld- knecht, N. J. Palleroni, J. W. Westley, and P. A. Miller, J. Anti* biotics :}6, 343 (1983).
6. Cf . : M. J. S. Dewar, The Molecular Orbital Theory of Organic Chem- istry (McGraw-Hill, New York, 1969), Chap. 8.
Rapid, On-Line Preconcentration, Matrix Normalization, and Signal Enhancement for Flame Atomic Absorption via Tubular Donnan Dialysis
J O H N A. K O R O P C H A K * and E W A D A B E K - Z L O T O R Z Y N S K A t Department of Chemistry and Biochemistry, Southern Illinois University, Carbondale, Illinois 62901
The application of tubular ion-exchange membranes for on-line use with flame atomic absorption (FAA) was investigated. Signal enhancement factors as high as 22 were observed within five minutes for a variety of cations. Enhancements improved with lower pH receiver solutions, in- creased tubing length, and increased temperature. Optimum receiver flow rates were a compromise between optima for independent Donnan di- alysis or FAA experiments. Low anion interferences were observed for phosphate-to-calcium mole ratios up to 80. Index Headings: Donnan dialysis; Atomic absorption spectrophotome- try.
INTRODUCTION
Donnan dialysis is a phenomenon which results in the transport of ions across an ion-exchange membrane un- der the influence of an ionic strength gradient. 1 If the volume of a high-ionic-strength receiver solution is sig- nificantly less than that of a sample solution, enrichment of sample ions in the receiver solution results. Analyti- cally useful enrichment factors as high as ~ 50 have been reported with reasonable sample and receiver volumes. 2 Further, counter ions are rejected; the type of membrane chosen (anion or cation exchange) determines which ions are preconcentrated. Membranes employed may be flat,
Received 9 April 1987. * Author to whom correspondence should be sent. t On leave from Department of Chemistry, Warsaw University, War-
saw, Poland.
for static experiments, ~ or tubular, for dynamic experi- ments. 4
Recently, the application of tubular-membrane Don- nan dialysis as an on-line preconcentrator of cations for atomic absorption spectrometry (AAS) has been report- ed) Receiver solution was drawn through the tubular membrane at the natural aspiration rate of the nebulizer for the AAS sample introduction system (typically 5-7 mL/min); the slope of a calibration curve was increased by a modest factor of 1.7.
Enrichment factors that can be achieved for tubular- flow Donnan dialysis are optimized at low receiver so- lution flow rates (<1 mL/min), since the apparent re- ceiver solution volume is decreased. 4 On the other hand, optimum sample flow rates for flame atomic spectrom- eters are significantly higher (5-10 mL/min), since the analyte mass flux to the flame is higher, providing higher signal levels. Clearly, the combination of the two exper- iments will require some compromise in sample flow rate to optimize system performance.
The combination of Donnan dialysis with flame atomic spectrometry has an additional advantage in that inter- ferences may be eliminated in a continuous fashion. For example, anions such as phosphate may depress signal levels for elements such as calcium by forming molecular species which are difficult to vaporize in many flames, s With a tubular cation-exchange membrane, calcium is preconcentrated in the receiver solution while the anions are rejected, eliminating the interferences. As an alter-
FIG. 1. Enrichment factors for 1-ppm Ca(II) sample collected Donnan dialysis (A) as a function of receiver flow rate of 0.1-M Sr(NOs)2/0.5- mM Al(NO3)JpH = 5 receiver; absorbance vs. flow rate for direct sample aspiration of 3-ppm Ca(II) to flame atomic absorption (0) ; air/ acetylene flame; 422.7-nm analytical wavelength.
native to Donnan dialysis, higher-temperature flames may be employed; however, ionization of analyte atoms may degrade optimum signals unless an ionization sup- pressor is added to the sample. 7 This process can be done in a continuous fashion with the use of Donnan dialysis through careful selection of the receiver electrolytes.
In this work we will describe a detailed parametric evaluation of the application of tubular Donnan dialysis to on-line use with flame AAS. Of particular interest was control of the receiver solution flow rate to determine a compromise between the relatively high sample flow rates, which optimize AAS, and the low flow rates, which op- timize enrichment via Donnan dialysis. Further, the elimination of classical anion interferences on cation analyses, the selection of receiver electrolytes to mini- mize ionization interferences, and the influence of other parameters, such as tubing length and temperature, are described.
EXPERIMENTAL
Static Donnan dialysis experiments were conducted for exploratory purposes with the use of a type P-1010 cation-exchange membrane (RAI Research Corporation, Hauppange, Long Island, NY) having a membrane area of 5.0 cm 2. Pretreatment procedures were identical to those described by Cox et al. s We pipetted the receiver electrolyte (5 mL) into the dialysis cell and then initiated dialysis by placing the membrane face in contact with the magnetically stirred 100-mL sample. After a specified dialysis time (usually 30 min), the receiver solution was transferred to a 10-mL volumetric flask, diluted to vol- ume, and analyzed for specified elements by atomic ab- sorption spectrophometry with the use of a Varian AA-475 (Palo Alto, CA).
Tubular Donnan dialysis experiments were conducted with the use of Nation 811 (Dupont Polymer Products Wilmington, DE) cation-exchange tubing (0.64-mm i.d. by 0.89-mm o.d.) of various lengths. Receiver solution was drawn through the dialysis tubing with the use of a Gilson (Middleton, WI) Minipuls II continuously vari- able peristaltic pump. Exact flow rates under specified
1.5
!
r~
)a-o-~.
\ ,
S J i i i i i i i 1 2 3 4 5 8
F ] OWr=~e. mL min m]
FIG. 2. Signal enhancement for on-line tubular Donnan dialysis as a function of flow rate; 0.1-M Sr(NO3)J0.5-mM AI(NO3)JpH = 5 re- ceiver; air/acetylene flame; 2-m tubing; 25 _+ 1°C; 1-ppm Ca(II); 422.7- nm analytical wavelength.
conditions were determined by volume measurements. During Donnan dialysis, the processed receiver solutions were either fed directly to the nebulizer and AAS, for absorbance measurement, or collected in an appropriate vessel, for later analysis. We stirred the sample solutions during the Donnan dialysis using a magnetic stirrer; a hotplate-stirrer was used for higher-temperature exper- iments.
The cation-exchange tubing was cylindrically coiled loosely around a three-pronged tubing holder, with fluid connections to the ends of the tubing from the receiver source reservoir and to the peristaltic pump. This assem- bly was placed into the vigorously stirred sample solution for Donnan dialysis. The length of the tubing required determined the dimensions of the tubing assembly. In order to maintain sample contact with the entire tubing length, we employed sample sizes for these studies that increased linearly from 100 mL for 1-m to 1000 mL for 10-m lengths.
Between measurements, the dialysis tubing was rinsed internally and externally with more concentrated re- ceiver solution (5 times higher), followed by water and finally by the receiver. This process typically required a total of about ten minutes and was adequate for routine work. For storage, we cleaned the ion-exchange tubing by pumping 1-M HN03 through the tubing while si- multaneously rinsing it externally.
The traditional means for evaluating the effectiveness of a Donnan dialysis experiment is the calculation of an enr ichment factor, which is the ratio of the analyte con- centration in the receiver after Donnan dialysis to the original sample concentration. In this work we were pri- marily interested in the improvements in signal that re- sult from the on-line Donnan dialysis experiment. Signal enhancement factors (SEF) reported here are the ratios of peak signals for on-line Donnan dialysis to the steady- state signal resulting from direct aspiration at the op- t imum FAA flow rate. Samples directly aspirated were either (1) standard solutions prepared in the receiver matrix to deconvolute the effects of the receiver matrix on signals, or (2) the standard solutions in a dilute acid matrix.
Elemental analyses were performed with the use of the
1232 Volume 41, Number 7, 1987
• 2 5
. 2
• o s r ' " .~..--"~
0 . . . . . . . . . . . , , , , , , , , i , i ,
1 2 3 4 5 6 7 8 g lO 11 12 Length of' tub tn 9. m
FIe.. 3. Signal vs. tub ing length for on-l ine D o n n a n dialysis us ing p H = 5 receiver (A) and pH = 1 receiver ( I ) ; 1 -ppm Ca(II) analyte.
T A B L E I. Signal enhancement factors for tubular Donnan dialysis- flame atomic absorption.
Analyte
Analyt i - cal wave-
l eng th (nm)
Signal e n h a n c e m e n t factors (SEF)"
Air /acetylene Ni t rous oxide/ f lame acetylene flame
A B A B
1.25 x 10 ' M Ca(II) 422.7 7.2 4.6 7.1 10.3 1.00 × 10-"> M Cu(II) 324.7 8.2 8.0 . . . . . . 1.00 × 10 "* M Co(II) 240.7 8.3 7.9 . . . . . . 2.4 × 10 6 M K(I) 766.5 7.6 10.8 . . . . . . 1.1 x 10 4 M Al(III) 309.3 4.5i . . . . 4.5 5.1
"A: SEFs calculated with respect to direct aspi ra t ion resul ts for s tan- da rds in receiver matr ix; B: SEFs calculated with respect to direct aspira t ion for s t anda rds in dil. HN03 matr ix; receiver, 0.1 M Sr(NO3)2 + 0.5 m M AI(NO3).~ a t pH = 1, ad jus ted with H N Q ; 5-m cat ion-exchange tubing; 25 _+ I°C.
t, La(NO:~)~ subs t i t u t ed for AI(NO~)3 in the receiver.
Varian AA-475, with the automatic gas control unit set at 13 L/min air and 1.6 L/min acetylene or at 13 L/min nitrous oxide and 4.5 L/min acetylene (reducing flames).
Standards and reagents were prepared from ACS re- agent-grade materials, except for the Sr(NO3)2 used in receiver solutions, which was Aldrich (Milwaukee, WI) Gold Label (99.999%). We prepared stock solutions of calcium by dissolving dried CaCO~ in dilute (3 % ) HNO3, while we prepared all other solutions from nitrate salts using dilute HNO3. Water used was deionized and dis- tilled.
RESULTS AND DISCUSSION
Factors relevant to the selection of a receiver solution for cation Donnan dialysis which result in high enrich- ment factors for a wide variety of cations independent of sample matrix are described elsewhere. 2 Simply, im- portant factors are the affinities of receiver cations for the exchange sites of the membrane and the valence of those cations; 2 for general purposes, a mixture of cations with a valence ranging from 1 to 3 is preferred. On the basis of these criteria, a commonly employed receiver solution consists of 0.2-M MgSO4 and 0.5-mM A12(SO4)~ at pH = 1.
For this work, additional requirements included the selection of receiver components which contained a cat- ion of low ionization potential to act as an internal ion- ization suppressant and the use of salts with NO3- as the counter ion to minimize vaporization interferences. As a result, the bulk of the data reported in this work was obtained with a receiver consisting of 0.2-M Sr(NO~)2 and 0.5-mM Al(NO3)3, adjusted to pH = 1 with HNOa. The choice of Sr(II) over Mg(II) was based on the sig- nificantly lower ionization potential of strontium. In stat- ic experiments, barium salts also provided adequate re- sults but produced lower enrichment factors (40 % lower) for Ca(II). In all respects but cost, La(III) salts were acceptable alternatives for Al(III). Aluminum is a va- porization interferent for Ca(II) but was not expected to be significant at the levels employed here. The selected receiver solution provided enrichment factors, as well as freedom from matrix effects, that were comparable to those reported for the MgSO4-AI2(SO4)3 system) Modi-
fication of the receiver composition may be subject to considerable variation based on the requirements of the analysis.
Figure 1 indicates the effect of flow rate on cation enrichment via tubular-flow Donnan dialysis where the receiver solution was collected for later analysis (A), and the effect of sample flow rate on absorbance for flame AA analysis (El). These data clearly indicate the need to compromise flow rates in order to optimize the on-line Donnan dialysis-FAA experiments.
Figure 2 indicates SEF values determined for the on- line Donnan dialysis experiment. In this case, these fac- tors were determined with respect to standards in the receiver matrix. Optimum signals were observed for 1.75 mL/min receiver flow. This value was employed for all further studies. These data were obtained at room tem- perature (25 - I°C), with a short length of dialysis tubing (2 m) and a receiver solution at pH = 5. Figure 3 indicates that significantly higher signal enhancement can be ob- tained with the use of greater lengths of tubing and lower- pH receiver solutions. The enhancement factors were independent of analyte concentration within experimen- tal error and within the upper limits of ionic strength for the Donnan dialysis experiment (0.01 M), as reported by Cox and DiNunzio2
Table I lists SEFs determined for a variety of elements with the use of air/acetylene and nitrous oxide/acetylene flames. SEFs calculated with reference to direct aspi- ration of standards in the receiver matrix (A) indicate that similar enrichment factors are obtained for the cat- ions tested; the one exception is the Al(III) data for which La(III) had to be substituted within the receiver. It should be stressed that these SEFs were obtained for room- temperature Donnan dialysis with 5 m of dialysis tubing.
As previously discussed, an additional feature of the Donnan dialysis experiment might be the inclusion of an easily ionizable component in the receiver to continu- ously suppress ionization interferences. In this case, the ionization potential for Sr(5.69 eV) is less than that for Ca(6.11 eV) and A1(5.98 eV). Ionization interferences are typically more severe for the hotter nitrous oxide/acet- ylene flame. Figure 4 is a plot of calibration data for the direct aspiration of calcium samples in the absence (A) or presence (0) of the receiver matrix. Clearly, strontium in the receiver solution is effective as an ionization buffer,
APPLIED SPECTROSCOPY 1233
. 35 , 2 5 ,m / Y
• 3 / / , /
• 25 / 2 " / = /
.~ i / " .
,45/ , 7'
• 0 5 ~ -
0 . . . . . . . . . . . , , , i . . . . , . . . .
. 5 1 1 . 5 2 2 . 5 C o , pp=
FIG. 4. Calibration data for on-line Donnan dialysis (O); direct as- piration of calcium in a dilute acid matrix (A); direct aspiration of calcium with the receiver as the matrix (e); nitrous oxide/acetylene flame; 422.7 nm.
providing significantly higher signals and essentially nor- malizing the matrix of the introduced sample. Also plot- ted are data for the on-line Donnan dialysis experiment (O), indicating the even greater enhancements which re- sult. The degrees of these effects are indicated in Table I by a comparison of SEFs calculated with the use of reference standards in the presence (A) or absence (B) of this receiver matrix. Higher B values indicate a matrix- induced enhancement, in addition to the Donnan dialysis enhancement.
Enhancements due to ionization suppression were ob- served for all elements with low ionization potentials (K, Ca, A1), especially with the hotter flame. The one excep- tion was Ca(II) with the air/acetylene flame. Similar sig- nal depressions occurred when La(III) replaced Al(III) in the receiver, or when only Ba(II) or Sr(II) were present as the major ionic strength contributors. This result sug- gests a general vaporization interference with the cooler flame for these high-ionic-strength samples.
Figure 5 indicates the times required to obtain max- imum response for the on-line Donnan dialysis experi- ment. At room temperature, typically 8 to 10 min were required for 5-m tubing. Lower times (5 min) were re- quired for shorter tubing lengths (1-2 m). At higher tem- peratures, the response time decreased significantly (~ 5 min at 65 _+ 1°C), and the signal enhancements were also increased. At 45 + 1°C, using a 10-m length of dialysis tubing, we determined signal enhancement factors in ex- cess of 22 for calcium. These effects are explained by increased cation transport rates at higher temperatures2 Clearly, further modification of the Donnan dialysis sys- tem to increase these mass transport rates should lead to greater enhancements and reduced response times. The slight decline in response after extended dialysis results from eventual depletion of analyte from the sam- ple solution.
Phosphate is a well-known interferent to the analysis of many cations by FAA, especially with air/acetylene flames. Figure 6 (O) indicates the extent of this inter- ference on Ca(II), even at low [PO4-q/[Ca +2] ratios for the direct aspiration of samples. With on-line Donnan dialysis at room temperature (A), this interference is
FIG. 5. Signal vs. time at 25 ± I°C (O), 45 + I°C (A), and 65 ± I°C (~); 1-ppm Ca(II) analyte.
eliminated up to [PQ-q / [Ca+q ratios of at least 40. At higher temperature (45 + l°C), the interference is small (< 10% ) up to mole ratios of 80. Simultaneous with the elimination of the interference, of course, is the same signal enhancement described earlier. The fact that higher temperatures provide low interference at higher [P04 a] suggests that the decreases that are observed may be due to solubility phenomena or the formation of non-ion- exchangeable calcium complexes. Further, at high mole ratios the ionic strength of the sample solution begins to approach the limits of the Donnan dialysis experiment.
CONCLUSIONS
Donnan dialysis using tubular ion-exchange mem- branes provides a convenient means of preconcentrating ionic species in low-to-moderate-ionic-strength samples in an on-line fashion for atomic spectrochemical analysis. Enhancements in signal over the direct aspiration anal- ysis as high as 22 were realized within 5 min in this work with the use of 10-m tubing lengths at 45°C with a 0.1-M Sr(NO3)J0.1-mM AI(NO~)JpH = 1 receiver solution. Higher values could probably be achieved with higher temperatures or greater lengths of tubing. Analyte cat-
.-2o \ "'~"~ "~'~ g-3o k.,..~.~ &-4n - ~ . .
~ -50 " ~
-so " ~ - ~ . . . ~
-')'0 , i i i i i , i 2Q 40 60 8Q IOO ]20 [ 40
[ PO,~3 / [ Co 3
FIG. 6. Effect of P04 '~ on signals for 2.0-ppm Ca via direct aspiration (O), or 0.5-ppm Ca via on-line Donnan dialysis at 25 ± I°C (A) and 45 ± I°C ((~); air/acetylene flame.
1234 Volume 41, Number 7, 1987
ions are concentrated in a normalized matrix (receiver), which can contain an ionization suppressant such as strontium to further enhance signals. Interferences due to sample anions are alleviated in this on-line fashion, and uncharged and high-molecular-weight species are excluded as well. These features should prove useful to trace analyses of dilute samples in the presence or ab- sence of interfering contaminants. The application of tubular Donnan dialysis to existing atomic spectrochem- ical instrumentation requires only the membrane tubing, a peristaltic pump, and a magnetic stirrer/heater. The simplicity of the approach should readily allow auto- mation and, perhaps, application to on-line monitoring of low-ionic-strength process streams.
Limitations of the approach include the upper limit to sample ionic strength for high enrichment, as well as trace impurities in the receiver solution. For example, trace Ca in Sr salts provided a significant blank for cal- cium studies. Further, the approach is sensitive to the chemical form of the element under study. This may
present problems for total element analyses of samples in which the analyte may exist in various forms (i.e., various cationic, anionic, or neutral forms) or it may be used to advantage for trace element speciation studies, s
ACKNOWLEDGMENTS We would like to thank J. A. Cox for helpful discussions and the
provision of ion-exchange membranes and tubing, and the Coal Re- search Center of SIU-C for partial support of E.D.-Z.
1. R. M. Wallace, Ind. Eng. Chem., Process Design Dev. 6, 423 (1967). 2. J. A. Cox, T. Gray, K. S. Yoon, Y. Kim, and Z. Twardowski, Analyst
109, 1603 (1984). 3. J. A. Cox and J. E. DiNunzio, Anal. Chem. 49, 1272 (1977). 4. J. A. Cox and Z. Twardowski, Anal. Chem. 52, 1503 (1980). 5. J. A. Cox and J. W. Carnahan, Appl. Spec. 35, 447 (1981). 6. R. Mavrodineanu and H. Boiteux, Flame Spectroscopy (John Wiley
and Sons, New York, 1965). 7. E. E. Pickett and S. R. Koirtyohann, Anal. Chem. 41, 28A (1969). 8. J. A. Cox, K. Slonawska, D. K. Gatchell, and A. G. Hiebert, Anal.
Chem. 56, 650 (1984).
Fluorescence Lifetime Study of Cyclodextrin Complexes of Substituted Naphthalenes
G R E G O R Y N E L S O N , G A B O R PATONAY, AND IS IAH M. W A R N E R * Department of Chemistry, Emory University, Atlanta, Georgia 30322
The interactions of a, 8, and 7-cyclodextrins and selected naphthalene derivatives as observed through fluorescence lifetime measurements are discussed in detail. These systems can be quickly characterized with the use of the parameters obtained from experimental fluorescence decay curves. The formation of inclusion complexes can be followed with the appearance of a long-lived fluorophore which contributes to the total fluorescence according to the cyclodextrin concentration. This fluoro- phore is determined to be an inclusion complex between a naphthalene and cyclodextrin. Index Headings: Fluorescence lifetime measurements; Cyelodextrins; Spectroscopic techniques.
INTRODUCTION
The study of cyclodextrin (CDx) complexes has been of general interest in the past several years because of their ability to form inclusion complexes with various organic solutes. 1 The unique torus shape of the CDx mol- ecule only allows appropriately shaped guest molecules to be included in the central cavity. These cavities dem- onstrate hydrophobic properties which provide favorable interactions for aqueous apolar solutes. 2 The three most commonly studied members of the CDx family, a-CDx, ~-CDx, and ~-CDx, have cavities with a diameter of 4.5- 6.0/~, 6.0-8.0/~, and 8.0-10.0/~, respectively.
Received 15 November 1986. * Author to whom correspondence should be sent.
Fluorescence lifetime measurements are especially useful in the study of CDx systems2 -s Both the absorp- tion and fluorescence bands of included polynuclear ar- omatic hydrocarbons are overlapping with the bands of the free fluorophore. In these cases, physical information from steady-state measurements about the interactions is not easily obtained at lo, CDx concentrations, where changes are subtle. Since the fluorescence lifetime is ex- tremely sensitive to a molecule's microenvironment, in- formation about the interaction between the included molecule and the CDx can be obtained for any CDx concentration. This is an advantage, since higher fluo- rophore:CDx complexes are observed in concentrated CDx solutions.
This paper demonstrates the differences observed in the cyclodextrin complexation behavior of structural iso- mers of naphthalene as characterized by fluorescence life- time measurements. Considerable interest has been shown in such complexation phenomena in the past several years. In chromatography, both cyclodextrin mobile phases and stationary phases have been studied to evaluate their utility for separating structural as well as enantiomeric isomers. 1°-12 Investigation of these complexation phe- nomena can provide useful insights into the interactions involved in cyclodextrin inclusion of a guest molecule. The effects of substitution on the fluorescence lifetime properties of the naphthalene : cyclodextrin complexes are presented and discussed.
