Anal. Chem. 1994,66,836-840
Evaluation of Some Immobilized Room-Temperature Phosphorescent Metal Chelates as Sensing Materials for Oxygen Yl-Mlng Llu, Rosarlo Perelro-Qarcia, Marla Jesh Valencla-Gonzhlez, Marta Elena Diaz-Garcfa, and Alfred0 Sant-Medel’ Department of Physical and Anaiyfcal Chemistry, Faculty of Chemistry, C/Julian Ciaveria, 8, University of Oviedo, 33006 Oviedo, Spain
8-Hydroxy-7-iodo-5quinolinesulfonic acid (ferron) chelates of some transition metals (including AI(III), Zr(IV), Ga(III), and Nb(V)), when retained on the surface of anion-exchange resin beads, exhibit strong phosphorescence at room temperature (RTP) in suspensions of aqueous and organic solutions or encapsulated in siliconerubber films. An oxygen transducer based on RTP quenchingmeasurementsof the AI-ferron chelate has been evaluated. The immobilized RTP metal chelate proved to be photochemically very stable, and its RTP emission is highly sensitive toward oxygen. A detection limit of 0.1%(v/ v) of oxygen in argon was found. Typical response times were less than 5 s for full signal change using naked sensing beads and ca. 20 s for gaseous mixtures and 2.5 min for solution samples when the sensing beads were embedded in a silicone film. The method has been successfully tested for dissolved oxygen determinations in tap waters. Research and development of optical sensors for oxygen measurements in gaseous mixtures, aqueous samples, and biological fluids are arousing great interest, particularly in relation to clinical and biological fields.1-3 Almost all optical sensors described so far for oxygen sensing are based on the measurement of the decrease in fluorescence intensity of a suitable indicator when it is quenched by molecular oxygen. Different fluorescence quenching based probes or indicators have been reported for sensing oxygen: polycyclic aromatic hydro~arbons,4*~ long-wavelength fluorescent indicators: and transition metal ~ h e l a t e s . ~ -Although ~ this last type of fluorescent probe exhibits attractive features, in principle phosphorescence quenching based oxygen sensors have several advantages over fluorescent ones;lOJl first, phosphorescence quenching is inherently far more effective by virtue of the longer lifetime; second, the analytical signal is a low-noise phosphorescent emission measured after any short-lived ( I ) Leiner, M. J. P. Anal. Chim. Acta 1991, 255, 209-222. (2) Wolfbeis, 0.S.;Leiner, M.J. P. SPIE Opr. Fibers Med. III 1988,906,4248. (3) Kaubky, H.; Hirsch A. Ber. Dtsch. Chem. Ges. 1931, 64, 2677-2686. (4) Peterson, J. I.; Fitzgerald, R. V.; Buckhold, D. K. Anal. Chem. 1984, 56,
62-67.
(5) Opitz, N.; Graf, H. J.; Luebbers, D. W. Sens. Actuators 1988.13, 159-163. (6) Wolfbeis, 0. S.;Carlini, F. M. A m / . Chim. Acta 1984, 160, 301-304. (7) Wolfbeis, 0.S.;Wcis, L. J.; Leincr, M. J. P.;Zieglcr, W. E. Anal. Chem. 1988. 60, 2028-2030. (8) Caraway, E. R.;Demas. J. N.; DcGraff, B. A.; Bacon, J. R. Anal. Chem. 1991.63, 337-342. (9) Li, X . M.; Ruan, F. Ch.; Wong, K. Y. Analyst 1993, 118, 289-292. (1 0) Huturbisc, R. J. Phosphorimerry. Theory, Instrumentation and Applications; VCH Publishers: New York, 1990. ( I I ) Vo-Dinh, T. Room Temperarure Phosphorimetry for Chemical Analysis; Wiley: New York, 1984.
858 A m l y t f c a l ~ b y Vd. , 66,No. 6, March 15, 1994
background luminescence has ceased, which should lead to much better detection limits for the phosphorescent sensors. Time discrimination (and so better selectivity) along with other characteristics tied to phosphorescence measurements, such as large singlet-triplet splittings, long excitation and emission wavelengths, etc., are also most preferable for optical sensing with today’s technology. In fact, the first optical sensor for oxygen determination described in the literature was based on room-temperature phosphorescence (RTP) quenching measurements.3 Grishaeva and Zakharov12 reported a series of studies on oxygen sensors based on the “after glow” quenching of some organic phosphors such as trypaflavine, acriflavine, or acridine, adsorbed on silica gel. Unfortunately, most of these phosphorescent organic luminophores are photolabileand, in some cases, completely moisture quenchable.lJ3 Recently, some RTP metal chelates have been exploited for oxygen sensing. Pd(I1)- (or Pt(I1)-) coprophorphyrins have been proposed for minute oxygen determinations, and this procedure has been extended for glucose measurements in biomedical samples.l6l6 Another very elegant oxygen optical sensor has been recently described, where the 02-quenchable RTP platinum dimer tetrakis(pyrophosphito)diplatinate(II) embedded in silicone rubber was used as the sensing material.” Previous experiments have demonstrated that 8-hydroxy7-iodo-5-quinolinesulfonicacid (ferron) forms negatively chargedchelateswith metalssuch as Al(II1) andNb(V) which emit phosphorescence at room temperature when the corresponding chelate is immobilized on anion-exchanger resins or in liquid solutions using “ordered media”.l”21 The phosphorescence emission of these complexes is strongly quenched by the presence of oxygen. In this paper we propose some ~~~
(12) Grishacva. T. 1.; Zakharov, A. I. Zh. Anal. Khim. 1990.45 (7), 1333-1337. (13) Lee, E. D.; Werner, T. C.; Seitz, W. R. Anal. Chem. 1987.59, 279-283. (14)Papkovskii, D. B.; Savibkii, A. P.; Yaropolov, A. I.; Ponomarev, G. V.; Rumyanwa, V. D.; Mironov, A. F. Blond. Sci. 1991, 2, 63-67. (15) Papkomkii D. B.; Savitakii, A. P.; Yaropolov, A. I. Zh. Anal. Khim. 1990,45
(7),
1441-1445.
( 16) Papkovskii D. 8.; Olah J.; Troyanowky I. V.; Sadmky N . A.; Rumyantcleva,
V. D.; Mironov, A. F.; Yaropolov, A. I.; Savitaky, A. P. Biasens. Btoeleciron. 1992, 7 , 199-206. (17) Li, X. M.;Won& K. Y. Anal. Chim. Acra 1992, 262, 27-32. (18) Pereiro Garcia, R.; Liu, Y. M.;Diaz Garcia, M. E.; Sanz-Medcl, A. A d . Chem. 1991,63, 1759-1763. (19) Sanz-Mcdd, A.; Martinez Garcia, P. L.; Diaz Garcia, M.E. A w l . Ckm. 1981,59,714-778. (20) Diaz Garcfa, M.E.; Fernllndez de la C a m p , M. R.; Hinze, W. L.; SanzMedel, A. Mikrochim. Acra 1-3, 269-282. (21) Liu, Y. M.; Garcia Aloruo, J. I.; Fernhdezde la C a m p , M. R.; DiazGarcia, M. E.; Sanz-Medel, A. Mikrochim. Acra 1991, I , 199-207.
ooo~~roo1e~1o~~6o~~o~.6o/o
ca lee4 m
RTP metal-ferron chelates of Al(III), Ga(III), Zr(IV), and Nb(V) immobilized on a strongly basic anion-exchange resin, as favorableindicator phases for oxygen sensing. The proposed indicators offer some distinct merits including favorable analytical wavelengths with excitation maximum around 400 nm and emission at 600 nm, a large singlet-triplet splitting of 200 nm and long triplet lifetimes (ca. 0.25 ms). The main characteristics of the RTP “active phases” for 0 2 sensing in gas mixtures, aqueous solutions, and organic solvents are described and the potential advantages of the proposed RTP oxygen sensors discussed. EXPERIMENTAL SECTION chemicals, The stronglybasic anion-exchangeresin Dowex 1 x 2 4 0 0 obtained from Sigma was cleaned thoroughly with 2 M HC1 and water before use, as described previously.18v22 Silicone rubber (Silicex, Silicona Hispania S.A.) was used as provided. All other chemicals were of analytical-reagent grade and they were used without further purification unless stated otherwise. Air (99.995%) and argon (99.998%) were purchased from Sociedad EspaAola del Oxigeno. Preparation of 0 2 Sensing Beads. For the preparation of the “active” Al-ferron-resin beads, the following procedure was employed: 0.3 mL of Al(II1) solution (1 g/mL), 35 mL M ferron (Fluka AG) solution, and 10 mL of pH of 3 X 1t3 5.5 acetic acid/sodium acetate buffer solution were mixed and diluted to 100 mL with water. The solution was pumped at a typical flow rate of 1 mL/min, using a FIA manifold, through a minicolumn packed with 0.25 mL of the resin.22 Then the resin was washed with 1 M NaCl solution and finally with distilledwater. These resin beads (RTP oxygen indicator) were kept in distilled water until use. Other metal-ferron complexes were prepared and bound to the resin in a fashion similar to that described for Al-ferron. Preparation of the Oxygen Sensing Films, The oxygen sensing films (ca. 50-pm thickness) were prepared by dispensing metal-ferron chelates bound to Dowex 1 X 2-400 resin particles in silicone rubber with a procedure similar to that described by Lippitsch et al.23 Silicone was selected because it is considered to be a most suitable material for oxygen sensing (high oxygen permeability, stability, absence of optical absorption bands, biocompatibility). The sensing beads were homogeneously mixed into the siliconeand rapidly spread on a quartz slide ( 1 mm thick, 6 mm wide, and 16 mm long) which was used as the membrane support. In order to allow an even film, membranes were covered for 2 h with a glass slide. The curing process was finished by letting the membranes stand in air at ambient temperature for another 2 h. Instrumentation. All RTP data were collected with a Perkin-Elmer LS 5 fluorescencespectrometer,which employs a xenon-pulsed (10-ps half-width, 50 Hz) excitation source and is equipped with a Perkin-Elmer 3600 data station. The delay time was normally set at 0.04 ms and the gate time was 2 ms for RTP measurements. Triplet lifetime measurements were made using the Obey-Decay application program. (22) Liu, Y. M.;Pereim Garcia, R.; Diaz Garcia, M.E.;Sam-Mdel, A. Anal. Chlm. Acru 1991,255, 245-251. (23) Lippitsch, M.E.; Pusterhofer,J.; Leiner, M.J. P.; Wolfbeis, 0.S.Anal. Chlm. Acru 1988,205,1-6.
I
m \z
! I
QUARTZ WINDOW
-
\
w‘
Flgure 1. Diagram of the Perspex flow cell (measurementsare @em in millimeters). ARGON from cylinder HUMIDIFIER VALVES AIR from cylinder
MFC SPECTROFLUORIMFTER
U
PERISTALTIC
PUMP
Solution Sample
Flgw2. Optosensingmnifold.(a,top)Fbwsystemforoxygensendng in gas samples; MFC, mss fbw controller. (b) Manifold for oxygen sensing In flowing liquid solutions.
For experiments with sensing beads, the beads were packed on a conventional Hellma flow cell (Model 176.52) as described elsewhere.18 In the case of experiments with sensing films, the membrane was positioned on a laboratory-made flowthrough cell with an inner volume of ca. 150 pL. Design and dimensions of this flow cell, which was fabricated from Perspex and fitted in the spectrometer sample compartment, are given in Figure 1. Optownsing Manifold and General Procedure. Figure 2 illustrates the two optosensing systems for introduction of gas mixtures and solution samples, respectively. For gases, by varying the relative flow rates of compressed air and argon, a steady environment with various concentrations of oxygen could be maintained in the flow cell. The oxygen concentration (% v/v) was calculated by dividing the air flow rate by the sum of the air and argon flow rates and considering that in Anal).dlcalChmWy, Vd. 66,No. 6, h4mh 15, l9M
83?
T a b 1. Chdcr ol MotaI-Forron Chdatr
IndlCator. chelate
Nb(V)-ferron Zr(IV)-ferron Al(II1)-ferron Ga(II1)-ferron
as tho Oxy+
maxX,(nm)
maxX,(nm)
Zh
ZdZh
365 380 390 390
570 585 800 600
45 38
6.3 5.8 5.5 5.2
86 60
a Z h and Z& are the RTP intensities when the sensor is exposed to argon and air, respectively.
wavelength, nm
-
Figurr 9. RTP spectra In the absence of oxygen: (left side) excitation spectra; (right side) phosphorescence spectra (a) Ai-ferron-CTAB solution [Ai] = 0.2 mg/L, [ferron] = 7.5 X lo-' M, pH 5.5. (b) Aiferron chetlate immobilized on Dowex 1x2-200 in water, pH 5.5. (c) AI-ferron chelate immobilizedon Dowex 1x2400 In sillconemembrane. (d) Blank (ferron adsorbed on anionic resin and embedded on silicone membrane). In cases (a) and (b), Na2S03was used as the oxygen scavenger.
pure air there is 21 -0%02.The total gas flow rate was kept constant at lOOmL/min. For liquids,water samples or organic solventswith different concentrationsof dissolved oxygen were prepared by mixing varying amounts of air-saturated liquid and argon-deoxygenated liquid in 1-L flasks. A peristaltic pump was used to generate a flow stream of the liquid sample continuously passing through the optical sensor. The flow rates of the samples were normally 1.5 mL/min. Considering the possible permeation of oxygen in air through the polymer tubings used in the peristaltic pump, the pump was positioned behind the sensor (see Figure 2b). All the phosphorescence intensities were measured at 600 nm with excitation at 390 nm, and all the measurements were made at room temperature (20 f 2 "C)and 101.325 kPa (1 atm).
RESULTS AND DISCUSSION Spectral Characteristics of Al(II1)-Ferron Chelates in Various Media. The RTP spectra of the complexes formed by Al(I1f) and ferron in different environments (a) cetyltrimethylammonium bromide (CTAB) micellar solution; (b) retained on Dowex 1 x 2 4 0 0 resin in an aqueous environment and (c) embedded sensing resin particles in a silicone membrane were recorded and the results are shown comparatively in Figure 3. A delay time of 0.04 ms was used for these experiments to ensure that any fluorescence from the complexes and the background had ceased. It should be pointed out here that the binary Al-ferron complex in solution is not phosphorescent at room temperature. As seen in Figure 3, the RTP luminophor in the three media showed almost identical spectra. This is not enough evidence to clearly demonstrate that the same species are responsible for the emission in the three environments. However, those results tend to indicate that the N(CHs)+ functional groups and the hydrophobic resin surface seem to provide protection for the 888 A?MlyikaIChembt~y,Vol. 66, No. 6, Merch 15, 1094
Al-ferron triplet state similar to that provided by the quaternary ammonium groups in the CTAB micelles.lOJ Using 0.04-ms delay time, the RTP blanks for the reagent (ferron) in CTAB solution, bound to the resin Dowex 1 X 2,181200reven in the silicone matrix (see Figure 3) arevirtually negligible. Choice of the Indicator Metal Chelate. As previously reported,*8-21 several transition metals form RTP complexes with ferron in micellar solutions under very similar chemical conditions and they exhibited similar spectroscopic characteristics. Using the same procedure, previously detailed for Al(III), the Nb(V)-, Zr(1V)-, and Ga(II1)-ferron chelates were immobilized on Dowex 1 X 2-400 resin particles. These particles were then embedded in a silicone layer spread onto a transparent quartz support. The RTP emission intensity, ZA~, and the IAr/zair ratio (where Zh and Zait are the RTP intensities measured when the membrane is exposed to argon and to air, respectively) were comparatively studied. A strong RTP emission may indicate a higher signal/noise ratio and consequently a superior 02 sensing sensitivity, but the ratio IAr/zair may be a more accurate method of evaluating the sensitivity of oxygen sensors.13 The experimental results observed in these experiments are given in Table 1. All of the four metal-ferron chelates tested proved to be phosphorescent and oxygen quenchable with different IAr/lair ratios. It is worth noting that the activated resin particles showed much higher Zh/Zair values when they stayed in water rather than in silicone membranes. As the Al(II1)-ferron chelate provided the highest RTP signal (while the ratios of zAr/zair were not substantially different for the different active surfaces studied), this chelate was selected for further experiments. Sensor Performance. The response time of the Al(II1)ferron sensing beads packed directly in the flow cell was less than 5 s for gaseous and liquid mixtures. When the sensing beads were embedded in the silicone film, Figure 4 shows a typical oxygen response of the Al( 111)-ferron sensor for gaseous mixtures. This response to gaseous oxygen was much faster than that observed for dissolved 0 2 in liquid mixtures, as shown experimentally in Figure 5 . The reversibility of the sensor was examined by exposing the device to argon and then to air (or to argon-deoxygenated water and air-saturated water) alternatively. The silicone sensor proved to be completely reversible for both, gaseous mixtures and water samples, as can be seen in Figure 5. Its response time changed, as expected, with the thickness of the sensing membrane and the flow rate of the samples. Typical response times for full signal change were in this case ca. 20 s for gaseous mixtures and ca. 2.5 min for solution samples, respectively, using the experimental conditions stated above.
'
7.5 %02
Aqueous'lsooctane Hexane Chlorofoim buffer
Flgurr 7. RTP emlsslon of senslng beads (AI(II1)-ferron on anbn exchanger) In different deoxygemted medie: aqueous buffer (0.2 M aoeticacidlsodlumacetate, pH5.5), Isooctane,hexane, and chloroform. IA = (I/Z-) x 100. I
T i m e scan
Flgwr 4. Typical response of the sensor slllcone membrane (wlth ANI1I)-ferron on anlonk exchanger as senslng phase) toward oxygen In gas mixtlwes Introduced In a contlnuous mode.
-
b
ARGON DEOXYGENATED WATER
-
Time scan Flgurr 1. Response-the curve and reverslbllity of the chosen oxygen
sensing membranetor determlnlng(a) gaseous samplesand (b) solutlon samples. 6
u 0
,
I
I
5
10
15
I 20
Oxygen concentration I
25
%I
Flgwr 6. Stern-Volmer plots of the quenchlng of Dowex 1x2-400 bindingAl-ferron chelates embedded In silicone rubber by oxygen. (A) RTP Intensity quenching; (0)RTP llfetlme quenching.
The effect of other gases such as carbon dioxide, helium, and nitrogen was evaluated for the proposed sensing beads and for the silicone sensing films. No interference of these gases was observed when they were introduced along with mixtures of pure oxygen and argon. Calibration experimentswere performed using conventional RTP intensity and lifetime measurements. Figure 6 shows the intensity and lifetime Stern-Volmer quenching plots. As can be seen, good linearity was obtained for both measurement modes. The slight differenceobserved experimentallybetween the slopes of both curves could be ascribed to a difference in Stern-Volmer constants obtained by each measurement method. When triplet lifetimes are measured, these constants
relate exclusively to dynamic phosphorescence quenching, while for the RTP intensites, the constant observed corresponds to the addition of static and dynamic deactivations. For 10 air to argon to air cycles, the relative standard deviation (RSD) for the RTP intensity in air was *2.0% and that in argon was fl%. The detection limit, estimated as the concentration of oxygen which produced an analytical signal equal to 3 times the RSD of the RTPintensity in argon, was 0.1%(v/v) oxygen in argon. Different experimentswere performed by measuring RTP signals with increasing/decreasing values of the partial pressure of oxygen. No hysteresis effects at all were observed. It should be pointed out that the sensitivity of this optosensor depends upon several important factors, mainly upon the particular metal-chelate used (e+ Al(III), Ga(III), Zr(IV), or Nb(V)), the type of polymer matrix, and the concentration of the indicator in the solid support.18 One or more of these factors could be modified in order to tailor the sensitivity for particular oxygen sensing purposes. It has been found that the RTP complexesimmobilized on the anion-exchange resin do not leach out either with water or with common organic solvents, such as ethanol, acetone, isooctane, or chloroform, at least during more than 3 months of use. It was observed that the selected sensing beads emit phosphorescence in the presenceof organic solvents. The RTP signals in these media after deoxygenation were even higher than in deoxygenated aqueous media, because of the known deleterious effect of water molecules on solid surface RTP.18 As can be seen from Figure 7, the RTP emission depends upon the organic solvent selected, with higher RTPs observed for the solvents with lower polarities. Figure 8 shows typical response curves of the sensing beads toward dissolved oxygen in isooctane. A final relevant property of the proposed sensor was its high photochemical stability in the presence of both oxygen and argon. No substantial decrease in RTP intensity of this active phase was observed after more than 20 h of continuous illumination. ApplicationtoRealSamples. The usefulness of the proposed RTP oxygen sensing silicone membrane was checked for the determination of the dissolved oxygen (DO) concentration in five different tap water samples following the general procedure. Argon-deoxygenatedand air-saturated waters, whose DO content was obtained from the atmospheric pressure and the tabulated data and further confirmed by the Winkler AnaWcal Chemistry, Vd. 66,No. 6,Wrch 15, 1994
018
Deownated Isooctane
Time scan (min.) Fbure 8. Response of the sensing beads toward dlssoived oxygen in isooctane. T a m 2. Comparison of the Proporrd RTP Oxygen Sensor wlth the Wlnkhr Method for Water-M#dved Oxygen (DO) Dotwmlnationr water DO found, pg/mL
samples
RTP sensor
Winkler’s method
I
2.08 4.68 6.67 7.19 8.25
2.23 4.75 6.42 7.40 8.34
I1 I11 IV V
method,24were used for sensor calibration. Five tap waters with different DO concentrations were analyzed with the proposed silicone film 02optosensor. To verify the reliability of the analytical values obtained from the RTP sensor, the same five tap water samples were analyzed by the Winkler method. The results obtained by the two methods are given in Table 2, which shows a good agreement between the values provided by the sensor and the classical method used as reference. (24) Standard Methods for the Examination Water and Wastewater, I5 4.; American Public Health Association, American Water Works Association, Water Pollution Control Federation: Washington, DC, 1965; pp 388-399. (25) Valencia, M. J.; Liu, Y. M.; DiazGarcia, M. E.;Sanz-Mcdel, A. Anal. Chim. Acta 1993, 283, 439-446.
040 Anahftlcal Chembtty, Vol. 66, No. 6, Mer& 15, 1994
CONCLUSIONS The proposed 02 optosensor offers some distinct merits as compared to many previously reported oxygen-sensitive indicators: one important aspect is the presence of appropriate functional groups in the reagent for ion-exchange immobilization on the resin. Stern-Volmer quenching constants can be reduced sometimes by 30-50% when the indicator is covalently bound onto a rigid support.’ However, the ferron chelates can be easily bound onto a solid surface via ionexchange processes, and interestingly, they phosphoresce at room temperature only when immobilized on strongly basic anion-exchange resins.18 Moreover, the immobilized ferron chelates possess high photochemical stability and they are not leached out by either aqueous or organic solvent solutions under normal conditions. In fact, the Al-ferron complex, when adsorbed onto the anionic exchanger, can be used for oxygen sensing in common organic solvents. These characteristics along with the sensor’s performance (high reproducibility, no hysteresis, good precision, long sensor lifetime) pave the way for an extensive field of applications. A simple flow injection analysis system based on this RTP oxygen transducer for submillimolar glucose determination in biomedical samples with a sampling frequency of ca. 40 h-l was recently developed.25 Other applications of this optosensor, particularly the development of biosensors in organic media, are currently under progress in our laboratory. ACKNOWLEDGMENT The financial support from “Fundaci6n para el Foment0 en Asturias de la Investigaci6n Cientlfica y Tkcnica” (FICYT) and “Fondo de Investigaciones de la Seguridad Social” (FISss) Project Ref. 90/0842 is gratefully acknowledged. Y.-M.L. thanks the Spanish Education and Science Ministry for a postdoctoral fellowship. Received for review August 3, 1993. Accepted December 2, 1993.’ Abstract published in Advance ACS Abstracts, January IS,
1994.
