BROAD RANGE pH SENSING NANOPARTICLES

FOR FIA AND SIA APPLICATIONS

Aleksandar Szchenyi
a
, Barna Kovcs
a,b



a
DDKKK Cooperative Research Centre Nonprofit Incorporated, PCS, Hungary

b
University of Pcs, Faculty of Natural Sciences, Institute of Chemistry, Department of General
and Physical Chemistry, PCS, 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 s 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 its 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
4C and the process was changed from acid catalyzed to base
catalyzed by addition of excess of NH
4
OH. The sol was then added
drop wise to mineral oil and stirred vigorously at 200C 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. 2024.
[3] Torsten Mayr, Ingo Klimant, Otto S. Wolfbeis, Tobias Werner, Analytica Chimica Acta 462 (2002) 110
[4] C.G. Niu, G.M. Zeng, L.X. Chen, G.L. Shen and R.Q. Yu, Analyst 129 (2004), pp. 2024.

Fig. 4. Calibration of sensing layer
Fig. 5. Phase shift as a functiom of modulation frequency
Acknowledgement: To Peter cs for thework on the
sythesis of thesensing materials and Ferenc Kaposvri
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.
-70,0
-60,0
-50,0
-40,0
-30,0
-20,0
-10,0
0,0
1000 10000 100000
Modulation Frequency (Hz)
P
h
a
s
e

s
h
i
f
t

(
d
e
g
)
APN/EtOH
Ru(dpp)3/EtOH + Ar
Ru(dpp)3+APN at pH 12
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 80C
Fig. 3. SEM images of the pH sensing nanospheres: Before thermal treatment (a,b) and after the thermal treatment (c,d)
a) b)
c)
d)
-18,00
-16,00
-14,00
-12,00
-10,00
-8,00
-6,00
-4,00
-2,00
0,00
0 1 2 3 4 5 6 7 8 9 10 11 12 13
pH
P
h
a
s
e

s
h
i
f
t

(
d
e
g
)
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.