Optical sensor for on-line determination of solvent mixtures based on a fluorescent solvent polarity probe

Sensors and Actuators B, 3 (1991) 267-272 267 Optical sensor for on-line determination of solvent mixtures based on a fluorescent solvent polarity probe zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCB Manfred A. Kessler and Jiirgen G. Gailer Anabtical Division, Institute of Organic Chemistry, Karl Franzens University, A-8010 Graz (Austria) Otto S. Wolfbeis Institute for Optical Sensors, Joanneurn Research, S~eyrer G. 17, A-8010 Graz (Austria) (Received March 5, 1991; accepted May 30, 1991) Abstract A new optical sensor for determination of solvent mixtures (such as water in organic solvent) is based on a fluorescent solvent polarity (SP) probe immobilized on an ion-exchange membrane. The fluorescent probe responds to changes in SP by both a shift in the fluorescence emission maximum and a change in fluorescence quantum yield. In this sensor, the analytical information is the relative fluorescence intensity measured at 620 nm (500 nm excitation). The sensor can be applied over a wide range of solvents. The response time (rW) is of the order of 15 s. Given the unique applications of fibre optic sensors (e.g., for remote sensing and sensing in explosive areas), the new sensor is expected to be applicable in process control. Introduction Continuous sensing of solvent polarity (SP) is of significant importance in process control when mixtures of solvents of different polarity are to be monitored. Because quite a variety of dyes is known to respond to SP by a change in their optical properties [l], it becomes obvious that such dyes may be exploited for sensing SP optically, particularly when combined with optical fibres onto which they may be immobilized. Optical parameters that may change with SP include absorbance, fluorescence intensity, fluorescence lifetime or efficiency of energy transfer from one chromophore to another [l]. Given the particular situation when using fibre optics, we prefer fluorescence over absorbance. Unfortunately, some of the common polarity indicators such as the ET dyes [2] are virtually non-fluorescent. On the other hand, most of the fluorescent polarity probes require UV or shortwave visible light excitation [3], which is not desirable when working with optical fibres with their poor transmissivity below 400-450 nm. Recently, we have reported on ketocyanine dyes 0925-4005/91/$3.50 that exhibit strong SP-dependent fluorescence [4]. The SP-induced shift of both the fluorescence excitation and the emission spectra makes it possible to sense SP optically using such probes. However, attempts to immobilize such dyes covalently on a solid support have been of little success so far. Electrostatic immobilization on an ion-exchange membrane appeared to be an attractive alternative, provided the dye could be furnished with more than one charged substituent in order to warrant strong electrostatic interaction with the solid support. We report here on a new ketocyanine dye possessing two (negatively charged) sulfo groups, the investigation of its SP-dependent fluorescence, its immobilization on an ionexchange matrix and its use for on-line determination of solvent mixtures, in particular water in ethanol. Experimental probes, chemicals and solvents The SP-sensitive probe used in this work was compound 4 (see Scheme l), which was prepared by the following four-step protocol: 0 1991 - Elsevier Sequoia, Lausanne 1-(3-(5-Sulfa-2,3-dihydro-IH-indol-I-yl)-2propenylidene-5-sulfo-2,3-dihydro-IH-indolI-ium perchlorate (3) This was prepared by analogy to a published procedure [4, 61 with modifications. Thus, a mixture of 2 (2.8 g, 14 mmol), tetramethoxypropane (1.13 g, 7 mmol), perchloric acid (70%; 2 ml) and anhydrous ethanol (5.5 ml) was heated to 60 “C for 10 min. After chilling, the obtained precipitate was sucked off to yield orange crystals (2.3 g, 60%). The material starts to decompose at 200 “C. IR spectrum (KBr): 1620, 1580 cm-‘. NMR spectrum (DMSO-d6): delta 8.98 (d, 2 H), 7.70 (s, 2 H), 7.46 (d, 2 H), 7.30 (d, 2 H), 5.90 (t, 1 H), 4.20 (t, 4 H), 3.27 (t, 4 H) ppm. . 2,5-Bis-[3-(5-sulfa-2,3-dihydro-IH-indol-Iyl)-2-propenylidenej-cyclopentanone (4) 4 Scheme 1. Synthetic pathway and chemical the fluorescent solvent polarity probe 4. structure of N-acetylindoline (1) This was prepared by dropwise addition of indoline (5.14 g, 43 mmol) in chloroform (25 ml) to a well-stirred solution of acetylchloride (3.35 g, 43 mmol) in chloroform (50 ml). The obtained precipitate was sucked off and dried to yield white crystals (3.4 g, 50%). M.p.: 218 “C. Indoline-Psulfonic acid (2) This was prepared according to a published procedure [5] with slight modifications: 1 (3.4 g, 21 mmol) was added in small portions to 8.8 ml of chilled chlorosulfonic acid under stirring. During the exothermic reaction, hydrogen chloride was evolved violently. Then, the mixture was kept at 50-60 “C for 1.5 h and finally poured onto 25 g crushed ice. The solution was filtered and kept in a refrigerator overnight. The precipitate was sucked off and washed with ice-cold water to yield colourless crystals (2.8 g, 65%) which, upon heating, start to decompose at 300 “C. IR spectrum (KBr): 2900, 2550, 1610, 1420, 1375 cm-‘. NMR spectrum (DMSO-d,): delta = 7.70 (d, 2 H), 7.47 (d, 1 H), 3.80 (t, 2 H), 3.25 (t, 2 H) PPm. This was prepared similarly to a previously published procedure [4]. Cyclopentanone (0.2 g, 2.4 mmol) and 3 (2.3 g, 4.3 mmol) were added to a previously prepared solution of sodium (0.6 g, 26 mmol) in anhydrous ethanol (13 ml). The reaction mixture was refluxed for 1 h and was finally of dark red colour. The precipitated product was sucked off and washed well with ethanol to yield the crude deep red dye (2.1 g). Since attempts to purify the dye by recrystallization or preparative TLC have not been successful so far, we applied it for sensing purposes without further purification. Ethanol/water mixtures (v/v) were prepared from 100% ethanol of extra pure quality (Merck, Dannstadt, F.R.G.) and double-distilled water. Preparation of the sensor membrane The sensing membranes used in this work have a 175 pm Mylar membrane as a solid support, onto which finely dispersed aminohexylcellulose (Serva, Heidelberg, FRG) was glued in a thin layer of typically 5-7 pm thickness using a hydrogel glue. The membrane was placed in the flow-through cell of the fluorometer and a concentrated aqueous solution of probe 4 was pumped through the cell, upon which binding of the dye to the surface of the membrane occurred. Instrumenta lion Uncorrected fluorescence excitation and emission spectra were obtained using an 269 Aminco SPF 500 spectrofluorometer equipped with a 250 W tungsten halogen lamp. Continuous monitoring of water/ ethanol mixtures was carried out using a Jasco 821 FP spectrofluorometer (Biolab, Vienna, Austria) equipped with a 150 W xenon lamp and a self-constructed 20 ~1 flow-through cell. Fluorescence was measured in the front-face geometry. The maximum spectral response was obtained by setting the excitation/emis408 588 zyxwvutsrqponmlkjihgfedcbaZYXWVUT 666 768 ml sion monochromators to 500 and 620 nm, welength Inn1 respectively. In order to decrease photoFig. 1. Fluorescence excitation and emission spectra of SP bleaching of the SP probe, the intensity of probe 4 in absolute ethanol (---) and in water (-). the exciting beam was reduced by a factor The spectral shift indicates a strong positive sokatochroof 10 using a violet Cokin Creative Filter (as mism. frequently applied in photography). In addition, scattered light from the sensor membrane was suppressed by the same filter, which acts as a 580 nm longpass filter. The response signal of the sensor was acquired and processed by a computer and recorded on a flatbed recorder. The whole system was under computer control using a Keithley 575 Measurement and Control System (Keithley Instruments, Vienna, Austria). All spectroscopic measurements have been carried out at 25 “C. zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA 618 ( 0 20 40 mter 60 80 100 in ethanol [vol. Xl Fig. 2. Near-linear relation between the wavelength of the emission maximum of the solvent polarity probe 4 and the Results and discussion zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA Design of the probe amount of water in ethanol. 4 was excited at its respective excitation maximum, ranging from 556 nm in ethanol to 577 nm in water. Quite a number of solvent polarity (SP) probes has been described in the literature. Table 1 gives a selection of dyes that have been considered as potential candidates for use in sensors. However, all of the dyes listed lack one of the following criteria: long-wave excitation, large Stokes’ shift, high fluorescence quantum yield, ease of immobilization and a strong dependence of optical properties on solvent polarity. We finally decided to use SP probe 4 for the design of an optical SP sensor. It has long-wave absorption and fluorescence, strong fluorescence in most organic solvents but poor fluorescence in water, and an acceptable Stokes’ shift. The synthetic pathway along with the chemical structure of probe 4 is given in Scheme 1. Due to the presence of two sulfa groups, the probe is readily soluble in polar solvents such as water and alcohols. Solutions exhibit strong yellow, orange, or red fluorescence depending on the respective SP. Fluorescence excitation and emission spectra in pure water and in ethanol are shown in Fig. 1. Both undergo long-wave shifts in changing from less polar to highly polar solvent (Table 2). We used this effect to design an optical sensor for solvent polarity, so as to measure the ratio of components in a binary mixture. We primarily focused on ethanol/water mixtures for practical reasons. Figure 2 shows that there is an almost linear relation between the emission peak wavelength of 4 and the water content in ethanol. In fact, the emission maxima may be used for determination of the ethanol/water ratio. For sensing purposes it is more practical, however, to work at fixed excitation and emission wavelengths. 270 TABLE 1. Solvent polarity probes and their photophysical properties Probe Excitation/emission maximum (in solvents of different polarity) Disadvantage Prodan” 350/452 (acetonitrile) 364/531 (water) requires UV excitation, oxygen quenches Nile red 485/525 (heptane) 530/605 (acetone) difficult to immobilize, water insoluble ET(30)b 795 (dioxane) 550 (ethanol) almost non-fluorescent 1-(4-Nitrophenyl)-6-phenylhexatriene 400/455 (hexane) 405/650 (methanol) requires UV excitation poor photostability “6-propionyl-2-dimethylaminonaphthalene. bData from ref. [l]. ‘Absorption maxima. TABLE 2. Spectral data of SP probe 4 in various solvents Solvent Polarity [ETN] Absorption/emission tnm) Water Methanol Ethanol 2-Propanol Dimethylsulfoxide Dimethylformamide 1.000 0.765 0.654 0.552 0.441 0.404 5771644 5551638 5561619 5201596 5201578 5321562 maximum Quantum yield (qualitative) low high high high high high a ETN values for solvent polarity are based on polarity indicator ET(30).In this scale, tetramethylsilane (TMS) is defined as the most apolar solvent (ETN=O.OOO),whereas water is the most polar one @TN= 1.000) (from ref. 1). Construction sensor and response function of the In order to obtain a sensor for continuous measurement of SP, we have immobilized probe 4 on a solid cellulosic support that is easily penetrated by solvent. A Mylar membrane is used as a mechanical support onto which we have immobilized aminohexylcellulose and the SP probe. Figure 3 shows the response of the sensor, operated at fixed wavelengths, to various ethanol/water mixtures. Unlike the case of correlation with the absorption maximum, the relation is non linear, simply because 4 exhibits much stronger fluorescence in ethanol than in water. Consequently, the highest sensitivity is achieved in the 80 to 100% ethanol range (Fig. 3). Figure 4 shows a typical response curve recorded from a series of mixtures containing small amounts of water in ethanol. Thus, a change from anhydrous ethanol to 95% ethanol causes a decrease in the signal by about one third. Given such a dramatic decrease in fluorescence intensity, a resolution of 0.5% water in >95% ethanol, and a detection limit of about 0.2% water in ethanol are realistic estimates. To the best of our knowledge, such a sensitivity of an optical SP sensor has not been reported so far. On the other hand, it is obvious from Fig. 3 that the sensor is rather insensitive in the 0 to 20% ethanol range, which is where the a.1 1 t t ethanol 100 X 0.0 1,. I,, 0 5 wter I. II 11, I,, 15 I,. I, ta tlm 25 Rid Fig. 3. Response time, relative signal change and reversibility of the SP sensor toward ethanol/water mixtures. The highest sensitivity obviously is observed in the 80 to 100% ethanol (v/v) range. 271 index changes. This would considerably increase the response time. In contrast to most chemical sensors and biosensors, the sensor reported here is a real sensor, in that it is based on a non-destructive physical process. It also may be applied to other solvent mixtures. 0 20 Conclusions * 180 48 64 zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA the kin1 Fig. 4. Response curve for ethanol/water mixtures. The sensitivity toward low levels of water is demonstrated by the a35 % decrease in fluorescence caused by 5 % water in ethanol (v/v). The signal drift indicates photobleaching that is caused by the strong xenon lamp of the fluorometer. alcohol content of most alcoholic beverages lies. The rather high sensitivity in the range from 80 to 100% ethanol is not so much due to the SP-induced spectral shifts in the absorption and emission maxima, but rather to the extreme SP dependence of the fluorescence quantum yield of 4. In this range, the sensitivity of the new sensor exceeds by far that of other sensors, such as those based on changes of the refractive index [7, 81. At present, some problems are encountered with the drifting signal of the sensor. This is assumed to be a result of photobleaching of 4 when it is exposed to the rather strong xenon lamp of the Jasco instrument. We expect to solve this problem by making use of a pulsed LED light source, which has considerably lower output power. It is desirable, however, to design an SP probe whose absorption has a better match with the emission of the green LED. An alternative way to overcome signal losses by photobleaching is to regenerate the sensor membrane by rinsing it with a stock of 4 in order to exchange the photodecomposition products. Leaching during operation does not occur to a measurable extent. The response time (t& is about 15 s, which is mainly caused by the dead volume of the flow-through cell. This is obvious, since there is no diffusional barrier and no chemical reaction step involved. However, for application to coloured or turbid solutions we recommend coating the sensitive membrane with an optical isolation layer, in order to exclude stray light and effects from refractive The sensor described here provides an easy means for on-line determination of low levels of water in ethanol. It is the first sensor that makes use of a fluorescent SP probe, and provides good sensitivity in the l-20% water in ethanol range and, more generally, for low levels of water in an organic solvent. Since the sensor does not require excitation with ultraviolet light, it is compatible with standard fibre optics and may be applied for remote process control. Potential fields of application include the monitoring of product quality in ethanol distilleries, on-line quality assurance during production of anhydrous ethanol, and on-line determination of water in technical solvents used in chemical plants. Acknowledgement We gratefully acknowledge the help of B. Weigl in software development. References Ch. Reichardt, Solvent and SoIvent Eficts in Organic Chetitty, VCH, Weinheim, 2nd edn., 1988, Ch. 6. M. A. Kessler and 0. S. Wolfbeis. ET(33), a solvatochromic polarity and micellar probe for neutral aqueous solutions, Cheer. Phys. Lipids, 50 (1989) 51. M. A. Kessler and 0. S. Wolfbeis, Polarity sensitive fluorescent probes, Appl. Fluoresc. Technol., 2 (1990) 11. M. A. Kessler and 0. S. Wolfheis, New highly zyxwvutsrqponml fluorescent ketocyanine polarity probes, S’ctrochim. Acfa, Part A, 47 (1991) 187. A. P. Terent’ev and M. N. Preobrazhenskaya, Introduction of substituents into the benzene ring of indole, Zh. Obshch. Khim., 30 (1960) 1218. D. Lloyd and H. McNab, Simple preparation of 2,3dihydro-lH-1,4-diaxepinium perchlorate, SynthesLs, (1973) 791. T. Takao and H. Hattori, Fluid observation with an optical fiber refractometer,Jpn. .I.Appl. Phys., 21(1982) 1509. 272 8 E. Smela and J. J. Santiago-Aviles, A versatile twisted Otto S. Woljbeis is professor of chemistry optical fiber sensor, Sensor and Actuators, 13 (1988) and head of the Analytical Division of the 117. zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA Biographies Manfred A. Kessler, born in 1964, received a Ph.D. in chemistry from Karl Franzens (KF) University in 1990. His major interests are in analytical chemistry, especially fluorescence methods, syntheses and applications of molecular probes, optical chemical sensors, flowinjection analysis (FIA), dye chemistry, solvent polarity, computer-controller data acquisition and programming in Turbo Pascal. Dr Kessler has authored several papers on new applications of molecular probes including determination of solvent polarity, cmc values of surfactants, and albumin in urine. Jiirgen G. Gailer, born in 1967, is an M.S.c. student at the Chemistry Department of KJ? University. His major interests are in analytical and organic chemistry. Institute of Organic Chemistry at the KF University in Graz, Austria, where he obtained his Ph.D. degree in 1972. He served as a post-doctoral fellow at the Max-Planck Institute for Radiation Chemistry in Miilheim (West Germany) from 1972 to 1974 and at the Technical University of Berlin in 1977. Dr Wolfbeis has authored more than 160 papers on optical sensors, fluorescence spectroscopy of plant natural products such as coumarins, flavons and alkaloids, and has edited a book entitled Fiber Optic Chemical Sensors and Biosensors. Other fields of research include three-dimensional fluorescence spectroscopy of biological liquids, and synthetic and spectroscopic work on fluorescent probes and indicators. He has given a number of invited lectures at international meetings and numerous guest lectures at universities and institutes. His current major research interests are in optical chemical sensors and biosensors.