A SO2-selective electrode based on a Zn

Analytica Chimica Acta 787 (2013) 57–63
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Analytica Chimica Acta
journal homepage: www.elsevier.com/locate/aca
A SO2 -selective electrode based on a Zn-porphyrin for wine analysis
˜ a , Maria
María Cuartero a , Célia G. Amorim b,∗ , Alberto N. Araújo b , Joaquín A. Ortuno
b
C.B.S.M. Montenegro
a
Department of Analytical Chemistry, Faculty of Chemistry, Regional Campus of International Excellence “Campus Mare Nostrum”, University of Murcia,
Murcia E-30100, Spain
b
REQUIMTE/Department of Chemical Sciences, Faculty of Pharmacy, University of Porto, R. Jorge Viterbo Ferreira 228, 4050-313 Porto, Portugal
h i g h l i g h t s
g r a p h i c a l
a b s t r a c t
• An innovative SO2 -selective electrode based on coordinated complexes between amines and metalloporphyrins.
• Lower detection limits allow the
evaluation of SO2 in different wine
preparation steps.
• Can be applied to the analysis of the
free and total SO2 in wine samples.
• Higher accuracy and precision when
compared with a reference method.
a r t i c l e
i n f o
Article history:
Received 10 April 2013
Received in revised form 17 May 2013
Accepted 21 May 2013
Available online 29 May 2013
Keywords:
Sulfur dioxide
Wine analysis
Metalloporphyrins
Potentiometry
Ion-selective electrodes
a b s t r a c t
This work describes the assessment of a SO2 -selective electrode based on the use of the neutral carrier 5,10,15,20-tetraphenyl(porphyrinate)zinc(II) in a PVC membrane plasticized with 2-nitrophenyl
phenyl ether. After being conditioned in 2 mol L−1 diethylamine solution for 24 h, the electrode exhibited
selective anionic response toward the analyte in a concentration interval of more than four decades,
with an slope of −59.5 mV dec−1 , a practical detection limit of 3.7 × 10−6 mol L−1 and a low limit of
linear range of 7.2 × 10−6 mol L−1 . The response mechanism is based on the displacement of the diethylamine:metalloporphyrin complex equilibrium within membrane bulk, inducing a variation in the
cationic-sites to ionophore ratio. In turn, free hydroxyl ions are complexed by the displaced ionophore
in a ratio 1:1 and translated as single negative charge nernstian response. Finally, the selectivity of the
electrode is evaluated in view of its application to wine analysis. Results had high accuracy and precision
when compared with a reference method.
© 2013 Elsevier B.V. All rights reserved.
1. Introduction
Sulfur dioxide (SO2 ) and other sulfites additives (E-220-228)
are commonly used as preservatives in food and beverages industries (Directive 95/2/EC, Directive 2006/52/EC). Particularly in wine
industry, SO2 is an ally of the producer since it prevents the development of undesirable natural processes in the winemaking process
which compromise the typical fruity flavor, color and freshness of
wines. Indeed, SO2 is added to must and juices both for a germicidal
∗ Corresponding author. Tel.: +351 918847877.
E-mail addresses: [email protected], [email protected] (C.G. Amorim).
0003-2670/$ – see front matter © 2013 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.aca.2013.05.038
action and for slowing down natural occurring oxidation processes
[1–3].
At the pH of wine (3–3.8), part of the SO2 added remains free
as bisulfite anion (HSO3 − ) and about 33–50% becomes chemically
combined with other native compounds. Sulfites act as antioxidants
by means of direct inhibition of polyphenoloxidases [4], hydrogen
peroxide removal and through reduction of quinones meantime
formed in the (bio)chemical oxidation processes [5]. The combined
fraction is presented either in reversible or as irreversible adducts
of aldehydes and ketones, due to its nucleophilic character. In part,
most of the bound SO2 can be released by proper heating or treating with a strong base. Nevertheless, the final amount present in
wines has to be strictly controlled since high levels of it produce
58
M. Cuartero et al. / Analytica Chimica Acta 787 (2013) 57–63
undesirable aromas and tastes and are also hazardous to human
health.
Each country has regulations that establish the limit of total SO2
permitted in wines which once more underlines the importance of
its control [6]. It would be convenient to have an expeditious technique that could be used to analyze the content of total and free SO2
in wines since the official method established by the European commission (EC) takes about 90 min per sample. For a simpler control,
Portugal suggested the Ripper procedure; nevertheless this still
takes about 30 min per sample [7]. Although the results obtained
by the reference method are more accurate than those obtained by
the Ripper method besides taking longer, it has the drawback of
requiring the operator to be extremely attentive because the temperature, gas flow and time steps need to be carefully controlled
[8].
Several methods have been published in the literature for the
determination of sulfite in wines. Worth of mention are the large
number of flow systems that have been proposed for this purpose, the main ones appearing in a review published in 2009 [4].
Generally, flow systems provide high sample throughput when
compared to conventional batch techniques, and when coupled to
electrochemical detection simple or none sample pre-treatment
is needed [9]. However, the acquisition costs are higher and due
to modular nature of the systems, require more skilled technicians. In recent years, voltammetric and amperometric (bio)sensors
[10–13], chemiluminescence [14] and ion chromatography [15,16]
methods have also been published for the SO2 determination,
although they usually only analyze the total SO2 and they are indirect and time-consuming methods. The proposal of potentiometric
methods is scarce and relies on the iodine oxidation by sulfite
[17,18]. Previous distillation or the use of gas permeation devices
was indispensable to improve selectivity relatively to other reducing compounds such as ascorbic acid in these methods.
Amines form coordinated complexes with metalloporphyrins in
reactions whose equilibrium could be shifted through addition of
strong nucleophiles [19]. Based on this a novel SO2 -selective electrode for the analysis of the free and total SO2 content of different
kinds of wine is presented. The principle of response was based on
the displacement of the complex formed between the diethylamine
and the lipophilic zinc(II)metalloporphyrin in the bulk of a permselective membrane. The developed electrode provides a simple,
economical and time-saving method which could be used by any
winemaker or controlling body.
2. Experimental
2.1. Reagents and solutions
All chemicals were of analytical reagent grade and Milli-Q water
was used throughout. Polyvinyl chloride (PVC) of high molecular weight (81392 Fluka), dioctyl sebacate (DOS) (CDS000559
Aldrich), 2-nitrophenyl phenyl ether (NPPE) (73307 Fluka),
2-fluoro-2-nitrodiphenyl ether (FNDPE) (47390 Fluka), tetradodecylammonium bromide (TDABr) (87249 Aldrich), potassium
tetrakis [3,5-bis-(trifluoromethyl)phenyl]borate (KTPB) (60588
Fluka), Araldite M (10951 Fluka), Ren HY 5162 (068620205 RenShape Solutions), graphite powder (<45 ␮m) (496596 Aldrich) and
tetrahydrofuran (THF) (186562 Sigma–Aldrich).
The ionophore 5,10,15,20-tetraphenyl(porphyrinate)zinc(II)
(ZnTPP) was synthesized from 5,10,15,20-tetraphenyl-21H,23Hporphirine (H2 TPP) (247367 Sigma–Aldrich).and ZnCl2 (8816
Merck).
Sodium metabisulfite (Na2 S2 O5 ), diethylamine (DEA),
hydrochloric acid (32%, w/w), sulfuric acid, chloroform,
dimethylformamide (DMF), n-octanol, 0.5 mol L−1 I2 solution,
starch soluble, acetaldehyde, sodium hydroxide (NaOH), 2-(Nmorpholino)ethanesulfonic sodium salt (MES), sodium chloride,
potassium chloride, magnesium chloride hexahydrate, copper(II)
chloride (CuCl2 ), calcium chloride dehydrate, ferrous chloride
tetrahydrated, sodium sulfite, sodium bicarbonate, sodium sulfate,
di-sodium tartrate dihydrate, sodium dihydrogen phosphate
anhydrous, sodium l-lactate, d,l-malic acid, histamine, tyramine,
cadaverine, putrescine, phenylethylamine, spermine, l-proline,
l-arginine, l-alanine and l-glutamine were also purchased from
Sigma–Aldrich.
A 0.1 mol L−1 Na2 S2 O5 standard solution was prepared by dissolving 0.19011 g of the commercial product in 100.0 mL of water.
Working solutions 0.01 and 0.001 mol L−1 were prepared by dilution of the standard solution. A MES buffer solution of 0.1 mol L−1
(pH = 5) and HCl/KCl solutions (pH = 1.6) at different KCl concentrations were also prepared. To obtain a 0.1 mol L−1 standard solution
of the corresponding sodium salt of malic acid, a 0.2 mol L−1 solution of the acid was potentiometrically titrated with 0.2 mol L−1
sodium hydroxide to end point. A 0.01 mol L−1 iodine standard
solution was obtained by dilution of the commercial solution.
2.2. Apparatus
Potentiometric measurements were made using a Crison
MicropH 2002 potentiometer (Crison Instruments, S.A., Barcelona,
Spain) and an Orion 90-00-02 Ag/AgCl double-junction electrode
(Orion Research, MA) with a 10% KNO3 solution in the outer compartment acting as reference electrode.
A spectrophotometer Jasco STR-707 V-660 and a pH-meter
Crison GLP22 equipped with a Phillips GAH 110 combined pH
glass electrode were namely used to characterize the synthesized ionophore 5,10,15,20-tetraphenyl(porphyrinate)zinc(II) and
for the measurement of solution pH.
2.3. Procedures
2.3.1. Synthesis of the ionophore
The ionophore Zn-TPP was synthesized via metallation of the
free porphyrin (H2 TPP) using ZnCl2 [20]. Both the H2 TPP (0.1 g) and
ZnCl2 (0.3 g) were dissolved in 10 mL of DMF and then refluxed at
153 ◦ C for 5 h. After the solution was cooled, DMF was completely
evaporated under vacuum (175 mbar) at 97 ◦ C in a rotary evaporator. The solid residue was dissolved in CHCl3 and purified with
a series of extractions in water. Afterwards, CHCl3 was evaporated
for several hours in cabinet and the remaining solid was allowed
to dry overnight. The resulting Zn-TPP was characterized by thin
layer chromatography and UV–vis spectrophotometry. The UV–vis
spectra revealed the maximum absorbance for H2 TPP and ZnTPP
at 417.8 and 421.8 nm, respectively. The obtained values were in
accordance with those reported in the literature [19,21]. Thin layer
chromatography was carried out in an n-octanol/CHCl3 (1:9) phase
to corroborate the purity of the synthesized compound.
2.3.2. Electrode construction
The electrodes used in the present work consisted of solid contact electrodes with the PVC membrane added drop wise on the
surface of a conducting solid support. To obtain this support a
mixture of Araldite M and HR hardener 1/0.4 (w/w) was homogeneously prepared. Briefly, 0.24 g of graphite powder was added to
0.2 g of the prepared mixture and the obtained paste was kneaded
to an appropriate homogeneity and consistency for incorporation
in the electrode body. The paste thus obtained was sufficient to
prepare about six electrodes. A portion of the paste was quickly
introduced in one side of a common Perspex tube of 6 mm of internal diameter, to a height of 0.5 cm, taking care not to leave pores
that might later have fluid leaks. On the opposite side a copper wire
M. Cuartero et al. / Analytica Chimica Acta 787 (2013) 57–63
Table 1
Compositions of the membranes assayed.
Membrane
Percentage (w/w) of components in the membrane
DOSa
A
B
C
D
E
F
a
b
c
NPPEb
FNDPEc
PVC
66
33
32.8
32.8
33
33
33
66
65.5
65.5
66
66
TDA-Br
0.5
1
K-TPB
Zn-TPP
0.5
1
1.2
1.2
established the electric contact with the graphite paste and, finally
the support was allowed to harden for 24 h. The surface of the support was then polished using sandpaper of different grit sizes to
provide an electrode surface with a specular gloss.
Membrane cocktails were prepared by dissolving 180 mg of PVC,
360 mg of the plasticizer (dioctyl sebacate (DOS), 2-nitrophenyl
phenyl ether (NPPE) or 2-fluoro-2-nitrodiphenyl ether (FNDPE)),
6 mg of the ionophore ZnTPP and 3 mg of ionic additive (KTCB or
TDABr) in 6 mL of THF to obtain membranes with the compositions shown in Table 1. Note that the membranes are not always
constituted by all the aforementioned components.
The corresponding cocktail was drop wised on the surface of
the support totally covering its area, and the THF was left to evaporate before the next membrane deposition. The cocktail drop was
repeated further six times and finally left to dry overnight before
conditioned in the corresponding conditions. When not in use, each
electrode was kept immersed in a solution similar to that used to
condition it.
2.3.3. General procedure for electrode calibration
All potentiometric measurements were carried out with continuous magnetic stirring and at room temperature.
When calibrating the electrode toward SO2 , the compound was
generated in situ by additions of Na2 S2 O5 to a medium of HCl/KCl
buffer at pH 1.6, following the reaction (Eq. (1)):
(1)
At a pH value of 1.6, according to the species distribution diagram, the 70% of the sulfur is in the SO2 form, a fact that has to be
taken into account when constructing the calibration graphs.
Serial calibration graphs of the electrodes evaluated were carried out by using micropipettes to add consecutively small volumes
of the corresponding solutions in 25.0 mL of the medium. The
steady-state potentials recorded were then plotted versus logarithmic values of the corresponding concentrations and the calibrations
parameters, slope (S) and detection limit (LD), were calculated by
fitting the calibration data to Eq. (2) [22]:
E = E 0 + S log(CSO2 + LD)
pH to ∼13. After 5 min the pH of the sample was lowered to 1.6 using
commercial HCl and the sample was diluted with HCl/KCl buffer in
a flask of 50 or 100 mL volume depending on the expected total
SO2 amount in the wine. The wine sample was then immediately
measured by the SO2 -selective electrode.
3. Results and discussion
1
1
Dioctyl sebacate.
2-Nitrophenyl phenyl ether.
Nitrodiphenyl ether.
2HCl + Na2 S2 O5 → 2SO2 + 2NaCl + H2 O
59
(2)
2.3.4. General procedure for the determination of free and total
SO2 in wines using the SO2 -selective electrode
For the determination of free SO2 , the standard addition method
was applied considering the sample treatment needed. About
30 mL of the sample were treated with concentrated HCl (32%) to
obtain a pH of 1.6. Just after, an aliquot of 25 mL was quickly measured potentiometrically by the electrode and suitable volumes of
Na2 S2 O5 solutions were successively added.
For the determination of total SO2 in wines the sample was first
treated to unbind combined SO2 . This was carried out according to
the method suggested by OIV [23]. A volume of 10.0 mL of wine was
alkalinized with 1.6 mL of 4 mol L−1 NaOH solution, adjusting the
As above described amines form coordinated complexes with
metalloporphyrins in reactions whose equilibrium could be shifted
through addition of strong nucleophiles. A good example of it is the
donor–acceptor reaction between the SO2 and diethylamine (DEA)
after the displacement of the last from its Zn-tetraphenylporphyrin
(ZnTPP) complex [19]. The Kassoc values of about 1.5 × 104 and
4.7 × 102 kg mol−1 for the complexes DEA:SO2 and DEA:ZnTPP in
chloroform, quantify the extent of this displacement which was
considered sufficiently sensitive to accomplish the colorimetric
determination of SO2 . However, the implementation of such simple
colorimetric procedure is problematic when dealing with colored
samples as it is the case of wines.
An alternative is to use the very same chemistry and to follow
potentiometrically the equilibrium of DEA:ZnTPP in polymeric
organic membranes exposed to different concentrations of SO2 . It
should be noted that several metalloporphyrins have been used
as neutral or charged carriers in the composition of membranes
for potentiometric sensors, successfully providing an improvement
of sensor performances regarding the sensitivity and selectivity
achieved [24]. Since the potentiometric behavior of the membrane
was a priori unknown, polymeric membranes plasticized with NPPE
of three different types were implemented: one containing solely
ZnTPP and the other two containing additionally an anionic or
a cationic additive (membranes A, B and C, Table 1). In the first
attempts, the incorporation of DEA in the membrane cocktail at
different molar ratios with respect to the incorporated amount of
ZnTPP was tried but the electrodes were irresponsive after immersion in solutions containing SO2 . A possible explanation is that the
cocktails medium used to prepare the membranes did not favor
the formation of the amine:metaloporphyrin complex fast enough
to avoid significant evaporation of the amine together with THF
during the polymerization of the PVC. Hence, it was assessed the
alternative incorporation of DEA in a conditioning process after the
electrodes construction. Therefore, the electrodes were soaked for
24 h in aqueous solutions of DEA with concentrations ranging from
0.01 to 3 mol L−1 . Noteworthy, the original dark red color of the
membranes changed to green when the DEA entered into the membrane and the complex was formed. This color change was evident
for electrodes constructed with the TDABr additive and for those
with membranes without any ionic additive, after being immersed
in DEA solutions with concentrations higher than 1 mol L−1 . The
electrodes with KTPB additive in the membrane composition did
not change noticeably the red color, which suggested a limited
diffusion of DEA into the membrane.
The calibration graphs, potential versus log[SO2 ], using the
implemented electrodes conditioned in DEA are shown in Fig. 1.
Two major features can be observed: one is that higher DEA concentrations rendered electrode responses shifted to more negative
potentials, and the other is the reduced response toward the
analyte presented by the membranes containing KTPB (Fig. 1a).
The membranes with TDABr and without ionic additive (Fig. 1b
and c) displayed slight potential increase to SO2 when conditioned in solutions of DEA below to 1 mol L−1 . However this last
electrode, containing only the carrier in the membrane and conditioned in the DEA 2 mol L−1 solution exhibited a nernstian negative
slope (-58.1 mV dec−1 ), a rather good practical detection limit
(3.9 × 10−6 mol L−1 ) and a lower limit of linear range of about
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M. Cuartero et al. / Analytica Chimica Acta 787 (2013) 57–63
Fig. 1. Calibration graphs for SO2 obtained with (a) membrane B; (b) membrane C and (c) membrane A, after conditioning the corresponding electrode for 24 h in 0.01 mol L−1
(), 0.1 mol L−1 (), 1 mol L−1 (), 2 mol L−1 (䊉) and 3 mol L−1 () DEA.
7.1 × 10−6 mol L−1 . The same type of electrode conditioned in DEA
3 mol L−1 , gave higher slopes but only for SO2 concentrations
above 10−4 mol L−1 . So the best conditioning was obtained in DEA
2 mol L−1 .
3.1. Rationale for the electrode response
The clues emerging from Fig. 1, help in the establishment of
the response mechanism of the electrode. First, taking into account
that membranes are immersed in water, the equilibrium of DEA in
aqueous solution (Eq. (3)) should be considered (pKa = 11.09 [25]):
DEA(aq) + H2 O ⇔
DEAH+
(aq)
+ OH−
(aq)
(3)
equilibrium above. The potential enabled by membranes containing anionic or cationic lipophilic additives differs only slightly
regardless the concentration of DEA in the solution used for conditioning (Fig. 1a and b). In fact, the equilibrium described above is
displaced to the left due to electrostatic repulsion by ions with the
same charge signal previously added during the preparation of the
membrane. For the membrane without any lipophilic additive, the
described effect is absent and so the potential of the electrodes is
strongly affected by the respective concentrations in the conditioning solution. For this membrane the heterogeneous co-extraction
constant is defined by Eq. (5):
Kcoex =
][OH− : ZnTPPn(org) ] aOH−(org)
[DEAH+
(org)
aDEAH+ aOH− [ZnTPP(org) ]n
aOH−
org
(aq)
After neglecting the effect of ionic strength for simplicity
reasons, the concentration values at equilibrium for both ions
on the right, increase approximately one decade (3 × 10−3 to
6 × 10−2 mol L−1 ) while the DEA(aq) increases almost three decades
(from 7.1 × 10−3 up to 2.94 mol L−1 ) in the conditioning solutions
used. Such great excess of DEA(aq) relative to the ions and a value
for its octanol–water partition coefficient (kow ) of 0.58 [26], reflect
in an increase in the transport into the bulk membrane due to the
diffusion gradient across the solution–membrane interface, where
it forms a coordinated complex with ZnTTP. This complex confers a green color to the membrane at ambient light, which is due
to the batochromic shift of about 15 nm in the Soret band of the
metalloporphyrin (absorption spectra not shown). Consequently,
the amount of ZnTTP that remains free in the membrane bulk is
drastically reduced.
Two different mechanisms should be considered for the entry
of DEAH+
ions into the membrane, beyond the entry due to its
(aq)
lipophilicity, the first is due to a cation-exchange with cations
associated with the anionic impurities present in the PVC and,
the second is promoted by coextraction together with OH− ions.
Considering these two phenomena, the following equilibrium can
be proposed according to the following equilibrium (Eq. (4)):
DEAH+
+ OH−
+ nZnTPP(org) ⇔ DEAH+
org
(aq)
(aq)
+ OH− : ZnTPPn(org) + OH−
org
(4)
The potential ranges observed for the three membranes at the
lowest SO2 concentration can be now described in light of the
= kDEAH+ kOH− ˇOH− :ZnTPP
(5)
n(org)
being ki the extraction constants into organic phase of DEAH+
and OH− and ˇ the cumulative binding constant for the complexes OH− :ZnTPP and OH− :ZnTPP2 in membrane bulk. The binding
constants of 3.9 × 105 and 3.1 × 103 kg mol−1 for the mentioned 1:1
and 1:2 complexes, respectively, depending of high or low cationicsite to free ionophore ratio, were reported in a recent work [27].
For a constant ZnTPP(org) in membrane, they are indicative that the
amount of complexed hydroxyl increases with the activity of both
ions in the conditioning solution. In accordance with phase boundary model [28], the developed potential in such membrane would
be predicted by Eq. (6):
EM = Econst + EPB = Econst
+
aOH−
RT
(org)
ln
F
aOH−
(6)
(aq)
where R, T and F are respectively the universal gas constant, the
absolute temperature and the Faraday constant, and aOH(org) the
activity of free hydroxyl ion in the organic phase. The term E const
includes the contribution of the reference electrode, the standard
potential of the selective membrane, the liquid junction potential
at the sample/bridge electrolyte interface and chemical potential
difference of the hydroxyl anion in both phases in standard conditions. From this model, it can be concluded that to the increase of
complexed hydroxyl ions corresponds a decrease of the free OH−
(org)
and so of EM potential. As shown in Fig. 1, the reason why for a constant SO2 concentration the potentials presented by membranes
M. Cuartero et al. / Analytica Chimica Acta 787 (2013) 57–63
61
decrease according the DEA amount in the conditioning solution is
now clear.
After being conditioned, the electrodes are immersed in a solution containing SO2 and this will displace the DEA:ZnTPP increasing
the amount of free metalloporphyrin. The amount of released metalloporphyrin is than able to react with free OH−
, thus causing a
(org)
potential decrease. For the electrode conditioned in DEA 2 mol L−1 ,
with
the amount of free ZnTPP newly formed complexes OH−
(org)
the stoichiometric ratio of 1:1, as evidenced by the nernstian slope
of the calibration curve obtained. For the electrode conditioned in
DEA 3 mol L−1 the amount of free metalloporphyrin is much lower
as result of shifted equilibrium with the excess of DEA(org) within
the membrane bulk. Thus the electrode does not change the potential until 10−4 mol L−1 of SO2 . The behavior of those electrodes can
now be summarized according to the sequence of equilibriums
presented in the two following equilibriums (Eqs. (7) and (8)):
SO2
DEA(aq) ⇔ DEA(org) + ZnTPP(org) ⇔ DEA : ZnTPP(org) −→DEA : SO2
+ ZnTPP(org)
ZnTPP(org) + OH−
⇔ OH− : ZnTPP(org)
(org)
(7)
(8)
The electrodes conditioned in the less concentrated solutions of
DEA (≤0.01 mol L−1 ) exhibit response profiles to SO2 very different.
The amount of the complex DEA:ZnTPP(org) is probably very low,
as well as DEAH+
and hydroxyl ions more strongly complexed
(org)
in 1:2 ratio with the metalloporphyrin. In the presence of SO2 an
increase of free metalloporphyrin is promoted which pulls back the
in DEA with consumption of OH−
. Under
conversion of DEAH+
(org)
(org)
these conditions, a potential increase is observed with increasing
SO2 concentrations.
Finally, the electrode initially immersed in the 1 mol L−1 of DEA
shows a bell shaped response as a consequence of both trends
explained before. The potentials generated by solutions containing SO2 up to the concentration of 3.2 × 10−5 mol L−1 show a slight
increase as explained for the electrodes conditioned in the less concentrated DEA solutions. Thereafter, the behavior becomes similar
to the electrodes presenting negative slope, but as the amount of
is limited, constant potential readings are observed for
free OH−
(org)
highest SO2 concentrations.
To evidence the fundamental role of the neutral carrier in
the selective response mechanism of the electrode toward SO2 , a
membrane containing PVC, NPPE and TDABr but without ZnTPP
(membrane D, Table 1) was tested either after or not the preliminary conditioning step with 2 mol L−1 DEA. As expected, SO2 did not
affect potential, because no reaction occurred between hydroxyl
ions and the neutral carrier. However, when the pH of the assay
conditions were raised from pH 1.6 up to 5 using a MES buffer solution, the electrode showed the typical anion response mechanism,
now due to the presence of bisulfite (Fig. 2). At the same time, the
membrane A tested in solutions at pH of 5 (at this pH value SO2
is quantitatively converted to bisulfite anion) exhibited constant
potentials only.
Fig. 2. Calibration graphs obtained using a membrane with PVC, NPPE and ZnTPP
(membrane A) conditioning in DEA 2 mol L−1 at pH 1.7 () and 5.0 () and conditioning in water at pH 1.7 (); and using a membrane with PVC, NPPE and TDABr
(membrane D) conditioning in DEA 2 mol L−1 at pH 1.7 () and 5.0 () and conditioning in water at pH 1.7 (䊉).
and the corresponding electrodes evaluated after being conditioned in 2 mol L−1 DEA for 4, 5, 17 and 24 h (Fig. 3). It was
found that an initial conditioning period of 24 h was necessary for
the three electrodes to provide nernstian anionic responses. With
more conditioning time the potentiometric response of the electrodes did not change. When the conditioning time was shorter,
insufficient DEA entered in the membrane and the responses
obtained were of the same type as when the electrodes were conditioned using less concentrated DEA solutions. Nernstian anionic
responses were obtained for the membranes plasticized with
DOS and NPPE (slopes of −57.3 and −59.5 mV dec−1 , respectively)
while the slope was lower with FNDPE (−50.5 mV dec−1 ). The
best PLD and LLLR values were obtained with NPPE (3.7 × 10−6
and 7.2 × 10−6 mol L−1 , respectively), followed by DOS (5.1 × 10−6
and 1.2 × 10−5 mol L−1 , respectively) and FNDPE (1.0 × 10−5 and
2.2 × 10−5 mol L−1 , respectively). Also, the membrane plasticized
with DOS had a higher drift (1.2 mV min−1 ) than those plasticized
with NPPE and FNDPE (0.2 and 0.3 mV min−1 , respectively). It was
supposed that potential drifts observed were due to DEA leaching
from the membrane into the sample solution since the membrane
changed its color from green to dark red with time. In accordance, it
was decided to follow further evaluations using only the electrodes
with membrane A, plasticized with NPPE, conditioned for 24 h in
2 mol L−1 DEA solution. Between measurements the electrode was
kept in the same solution in order to avoid leaching or evaporation.
3.3. Characteristics of the electrode response
3.2. Selection of the plasticizer
It is well known that potentiometric characteristics of ionselective electrodes are influenced by the nature of the membrane
plasticizer, namely by affecting the solubility and mobility of
ions and molecules within membrane bulk and the leakage of
membrane components into the sample matrix [28]. Hence, three
PVC membranes containing ZnTPP and plasticizers with increasing dielectric constant DOS (ε = 4.0), NPPE (ε = 24.0) or FNDPE
(ε = 50.0) (membranes E, A and F, respectively) were implemented
Calibration parameters of the developed SO2 -selective electrode, slope and practical detection limit, together with other
important performances characteristics, are shown in Table 2. The
response time of the electrode was taken as the time needed to
reach the steady-state potential ±0.1 mV for the lowest and the
highest concentrations corresponding to its linear range. Repeatability of the electrode was evaluated by making four successive
calibrations in the same day and conditioning the electrode in
the DEA solution between calibrations for about 20 min until
62
M. Cuartero et al. / Analytica Chimica Acta 787 (2013) 57–63
Fig. 3. Calibration graphs for SO2 obtained with (a) membrane E; (b) membrane A and (c) membrane F, after conditioning the corresponding electrode in DEA 2 mol L−1 for
4 (), 5 (), 17 () and 24 h (䊉).
the original potential baseline was restored (±5 mV). This reconditioning step is necessary to obtain a good repeatability of the
electrode response and if not carry out, the green color of the membranes is lost in consecutive calibrations as reflected by a loss of
the potentiometric response of the electrode. The long-term stability was studied by recording the response of an electrode over
one month. Visually, the membrane changed its initial appearance when the electrode was used repeatedly. After 30 days, the
membrane became opaque and showed inhomogeneities like little bubbles and roughness in the surface. Finally, the membrane
becomes bicolor, being the center green-brown and the borders
green. After more than 1 month, the membrane started to separate from the electrode body. However, the electrode could be
used in continuously work for 15 days without appreciable variation in the calibration parameters. Finally, the responses displayed
by six different electrodes, all of them constructed with the same
membrane cocktail, were evaluated for the between-electrode
intraday repeatability. The results obtained are also enabled
in Table 2.
Taking into account all the above mentioned characteristics, it can be said that the developed SO2 -electrode provided
a reproducible nernstian response above a SO2 concentration of about 6 × 10−6 mol L−1 , with a low detection limit
of 3.2 × 10−6 mol L−1 . The response time was lower than 20 s
Table 2
Response characteristics of the developed SO2 -selective electrode.
Parameter
Value
Slope (mV dec−1 )a
Practical limit of detection (mol L−1 )a
Lower limit of the linear range (mol L−1 )
Response time (s)b
Long-term stability
Slope (mV dec−1 )a
Practical limit of detection (mol L−1 )a
Lower limit of the linear range (mol L−1 )c
Intra-day repeatability
Slope (mV dec−1 )c
Practical limit of detection (mol L−1 )c
Lower limit of the linear range (mol L−1 )c
59.8 ± 2.5
(3.7 ± 0.8) × 10−6
(8.4 ± 1.4) × 10−6
<20
a
b
c
58.8 ± 1.3
(3.2 ± 0.5) × 10−6
(6.0 ± 0.7) × 10−6
57.5 ± 1.5
(4.3 ± 1.9) × 10−6
(9.8 ± 0.1) × 10−6
Mean ± SD (n = 4).
Corresponding to SO2 concentration range 1 × 10−6 to 1 × 10−3 mol L−1 .
Mean ± SD (n = 7) over 15 days.
and the electrode worked for at least 15 days in continuous
operation.
3.4. Interferences
The response of the developed electrode was evaluated toward
the main cations and anions present in wines, in their chloride and sodium forms, respectively, and also toward the amino
acids and biogenic amines, which are prone to enter into the
membrane by forming a complex with ZnTPP. Proline, arginine,
glutamine and alanine are the main amino acids present in wines
and their common levels are well known, while putrescine, cadaverine, spermine, histamine, tyramine and phenylethylamine are
biogenic amines that may be present in wines. Although there
are no definitive regulations regarding the concentration of biogenic amines in wine, some countries have set histamine import
limits and Office international de la vigne et du vin (OIV) recommendations state that the histamine content should not exceed
12 mg L−1 .
The separate solution method [29,30] was used to test the selectivity of the electrode toward several cations and anions (Na+ , Mg2+ ,
Cu2+ , Ca2+ , HCO3 − , SO4 2− , H2 PO4 − , malate, tartrate and lactate).
This method was selected since the ion does not induce an output
of DEA from the membrane and so the re-conditioning of the electrode was not necessary. No selectivity coefficients were calculated
for these compounds since the electrode displayed no response for
any of them. In the case of K+ , since KCl is a component of the buffer
used for the medium, the fixed interference method was used by
making calibrations for SO2 in HCl (at pH 1.6) in presence of 0.2,
0.02 and 0.002 mol L−1 KCl. The presence of 0.2 mol L−1 KCl favors
the response of the electrode toward SO2 at the higher SO2 concentrations since it maintains the ionic strength of the medium
constant.
To evaluate the influence of amino acids and biogenic amines,
the amount at which the base line of the electrode varies in 10 mV as
a consequence of the entry of the nitrogenous compound into the
membrane through complexation with ZnTPP was obtained. The
results obtained, together with the amount of each amino acid normally present in wines are shown in Table 3. For biogenic amines,
a value of 12 mg L−1 was used as reference. As can be seen, all the
compounds tested lowered the potential baseline of the electrode
in approximately 10 mV when the amounts largely exceeded the
normal stated values.
M. Cuartero et al. / Analytica Chimica Acta 787 (2013) 57–63
Table 3
Interference studies.
Compound
Approximate content
in wine (mmol L−1 )
Amount from which the
potential base line varies in
10 mV (mmol L−1 )
Proline
Arginine
Alanine
Glutamine
Histamine
Tyramine
Cadaverine
Putrescine
Phenylethylamine
Spermine
45
1
2
1
0.065
0.069
0.069
0.075
0.076
0.035
1000
200
1000
200
320
280
400
620
630
460
1
2
3
4
5
6
a
b
Free SO2 (mg L−1 )
Total SO2 (mg L−1 )
ISEa
ISEa
16.70
63.23
11.76
21.53
32.70
18.74
Referenceb
±
±
±
±
±
±
0.49
3.12
0.84
0.66
2.58
0.58
17.61
59.41
10.82
20.10
34.95
17.75
±
±
±
±
±
±
2.30
2.43
1.27
3.17
2.17
1.0
129.87
147.74
102.78
112.08
147.86
125.81
Referenceb
±
±
±
±
±
±
4.55
3.19
3.74
4.62
3.36
3.01
127.90
150.69
109.43
111.51
141.41
133.08
±
±
±
±
±
±
3.40
2.65
1.27
2.14
3.38
0.62
Mean value ± SD of two determinations made with three electrodes.
Mean value ± SD of two determinations.
The usefulness of ion-selective electrodes in the analysis of interfering colored matrices as is the case of wines,
is here evidenced in the determination of SO2 . The pair
diethylamine:zinc(II)-metalloporphyrin as the main membrane
component of the proposed SO2 electrode ensured an anionic nernstian response and enabled a selectivity toward the analyte. The
quality of results furnished was statistically similar to the reference procedure commonly used. Simplified sample pretreatments,
reduced analytical costs and time of analysis were here evident in
an easiest implemented procedure with increased physical ruggedness. Point out that potentiometric analysis of sulfur dioxide is a
good alternative to usual control procedures adopted in wineries.
Acknowledgements
M.C. and J.A.O. gratefully acknowledge the financial support of
the Ministerio de Ciencia y Tecnología, Spain (project CTQ201127049). M.C. is grateful to the University of Murcia for a grant.
3.5. Determination of SO2 in wines
The developed SO2 -electrode was applied to the analysis of the
free and total SO2 in table wines using the process suggested in
Section 2. The results are shown in Tables 4 and 5. The results
obtained for white wines were compared with those enabled by
the Ripper method. In the case of red wines, the Ripper method
provides dispersive results, since wine color interferes with the end
point detection of the titration, so recovery studies were performed
Table 5
Results obtained for the determination of free and total SO2 in red wines.
Red wine
sample
Free SO2 a (mg L−1 )
Added
(mg L−1 )
Found (mg L−1 )
Recovery (%)
1
14.17 ± 0.49
25.6
51.3
76.9
26.8 ± 1.3
49.0 ± 1.5
76.6 ± 0.9
104.7 ± 4.4
95.5 ± 3.0
99.6 ± 1.2
2
8.52 ± 0.31
25.6
51.3
76.9
25.3 ± 1.1
51.2 ± 0.9
76.1 ± 1.2
98.7 ± 4.1
99.9 ± 1.8
98.9 ± 1.6
3
14.30 ± 0.97
25.6
51.3
76.9
25.9 ± 0.7
51.0 ± 0.6
76.8 ± 1.8
101.1 ± 2.6
99.4 ± 1.2
99.8 ± 2.4
Red wine
sample
Total SO2 a
Added
(mg L−1 )
Found (mg L−1 )
Recovery (%)
1
124.03 ± 3.5
256.4
512.8
769.2
259.3 ± 6.5
518.9 ± 8.5
765.2 ± 7.1
101.1 ± 2.5
101.2 ± 1.7
99.5 ± 0.9
2
132.28 ± 4.87
128.0
256.4
384.6
129.3 ± 3.5
258.3 ± 2.8
390.1 ± 3.3
101.0 ± 2.7
100.8 ± 1.1
101.4 ± 0.9
3
92.31 ± 2.48
128.0
256.4
384.6
129.9 ± 2.8
257.4 ± 2.4
383.6 ± 3.0
101.5 ± 2.2
100.4 ± 0.9
99.8 ± 0.8
a
instead. In all cases the recovery results ranged from 95.5 to 104.7,
and in general were quite close to 100%.
The results obtained by Ripper and the proposed method
when analyzing white wines of 2011, from different demarked
regions of Portugal (Douro, Alentejo, Península de Setúbal, Vinhos
Verdes/Minho), were compared by applying the t-test and the Ftest at 95% confidence level. The calculated F and t values did not
exceed the theoretical values (F = 15.10; t = 2.45), indicating that no
significant difference in accuracy or precision existed between the
two methods.
4. Conclusions
Table 4
Results obtained for the determination of free and total SO2 in white wines.
White wine
sample
63
Mean value ± SD of two determinations made with three electrodes.
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