BROAD RANGE pH SENSING NANOPARTICLES FOR FIA AND SIA APPLICATIONS

Aleksandar Széchenyi a, Barna Kovács a,b

a DDKKK Cooperative Research Centre Nonprofit Incorporated, PÉCS, Hungary b University of Pécs, Faculty of Natural Sciences, Institute of Chemistry, Department of General

and Physical Chemistry, PÉCS, Hungary

Introduction: One of the major advantages of optical sensors is that they are usually made of a sensor cocktail that could be cast on any optically transparent support. Depending on the requirements of the application this way sensors having different geometry could be obtained, such as films, spots, tubes. When the measurements should be made in a flowing stream, planar sensors mounted in a flow-through-cell or layers prepared on the inner wall of a capillary are advantageous. However, these sensor formats allow the measurement of a single parameter of the sample. By injecting different sensors beads into the sample/carrier stream, different analytes could be measured by using the same light path. In the present work the use of fluorescent, pH sensitive nanoparticles in different flow injection setups were tested. For this purpose core-shell type silica nanosphere were synthesized using a sol-gel technique. To eliminate the common fluorescence intensity measurement interferences, we have used a dual lifetime referencing (DLR) method [1]. It is a principle to reference fluorescence intensities via fluorescence decay times. The DLR method uses two fluorescent dyes with overlapping spectroscopic properties, one pH-sensitive, short-lived indicator and a pH-insensitive reference dye with a decay time in the μs or ms range. N-allyl-4-piperazinyl-1,8-naphthalimide (APN) have been used as fluorescent pH indicator [2] . Ruthenium(II) tris(diphenylphenanthroline) (Ru(dpp)3) complex [3] has been used as a reference fluorophore because of it’s high optical quantum yield and high thermal stability.

Experimental: The synthesis of APN was similar to that reported by Niu et al. [2]. The pH sensing principle of APN is shown in Fig 1. and its fluorescence properties on Fig 2. The core of the silica sphere was prepared by dissolving an appropriate amount of Ru(dpp)3 in the tetraethoxysilane (TEOS), after the addition of acid catalyst, ethanol and deionized water the mixture was stirred to form acid catalyzed sol. The sol formation was allowed to proceed for one hour, then the mixture was cooled to 4°C and the process was changed from acid catalyzed to base catalyzed by addition of excess of NH4OH. The sol was then added drop wise to mineral oil and stirred vigorously at 200°C until silica spheres appeared. The spheres were filtered, washed with ethanol and deionized water. Cocktail for the shell of the sensor was prepared in to phases. For preventing the leaching of pH sensing dye, APN was covalently bond the to sol-gel precursor vinyltriethoxysilane (VTES) by irradiating their mixture (molar ratio 1:3) with UV lamp (366nm) for 30 minutes. TEOS, ethanol, water and HCl were added, sonicated for 5 minutes and left to form gel for 1 hour. The shell was formed by adding the Ru(dpp) containing spheres to shell forming gel and stirred for one hour at room temperature. The silica spheres was filtered in the centrifuge with 0,22 mm pore diameter filter and washed with ethanol to remove the unreacted components. The resulted particles were dispersed in the ethanol until use. The Phase shift measurements were performed with dual-phase lock-in amplifier (DSP830, Stanford Research inc.) in home made flow through cell. Optical system consisted from blue led (430 nm) with band pass filter, bifurcated fiber bundle, and Hamamatsu (H5783-01) PMT with long pass filter (550 nm).

[1] I. Klimant, C. Huber, G. Liebsch, G. Neurauter, A. Stangelmayer, O. S. Wolfbeis, New Trends in Fluorescence Spectroscopy, Springer Series on Fluorescence, 2001, 257-274.[2] C.G. Niu, G.M. Zeng, L.X. Chen, G.L. Shen and R.Q. Yu, Analyst 129 (2004), pp. 20–24. [3] Torsten Mayr, Ingo Klimant, Otto S. Wolfbeis, Tobias Werner, Analytica Chimica Acta 462 (2002) 1–10[4] C.G. Niu, G.M. Zeng, L.X. Chen, G.L. Shen and R.Q. Yu, Analyst 129 (2004), pp. 20–24.

Fig. 4. Calibration of sensing layer

Fig. 5. Phase shift as a functiom of modulation frequency

Acknowledgement: To Peter Ács for the work on the sythesis of the sensing materials and Ferenc Kaposvári for SEM images

 

Phase shift measurements

Fig. 2. APN fluorescence on different pH

Fig 1. Fluorescence enhancement mechanism

Calibration Results: SEM images of the synthesized nanospheres are shown in the Fig 3.. The main size of the particles was 275 and 380 nm respectively. The thermal treatment of the nanospheres did not effect the morphology, the particles formed a stabile suspension in ethanol. Proton strongly enhance the fluorescence intensity of APN, which shows no fluorescence above pH 12. The phase shift of the reference material and the overall sensor particles as a function of the modulation frequency is shown in the Fig 5. It has been found that the difference in the phase shift have a maximal value at the 10 kHz modulation frequency. Fig. 4. shows the calibration curve of the sensing nanospheres.

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Ru(dpp)3+APN at pH 2

Ru(dpp)3+APN in DI water

Ru(dpp)3/TEOS+Ar 200 °C 24h

Ru(dpp)3/TEOS+Air dry at 80°C

Fig. 3. SEM images of the pH sensing nanospheres: Before thermal treatment (a,b) and after the thermal treatment (c,d)

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c) d)

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Conclusion: A pH sensing nanospheres has been prepared with covalently bonded pH sensitive dye and co immobilized reference dye. The sensor calibration have a liner correlation in the pH range 7-11,5. Further investigation is needed for pretreatment and conditioning of the sensing nano spheres to improve their performance in the pH sensing. The examined combination of the materials shows promising results for the further development of pH sensor.