Food Chemistry 134 (2012) 483–487

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Food Chemistry
journal homepage: www.elsevier.com/locate/foodchem

Analytical Methods

Development of a sensitive potentiometric sensor for determination of fumaric

acid in powdered food products
Alberto Oppermann Santini, Helena Redigolo Pezza, Leonardo Pezza ⇑
Instituto de Química, São Paulo State University – UNESP, P.O. Box 355, CEP 14801-970, Araraquara, SP, Brazil

a r t i c l e i n f o a b s t r a c t

Article history: A new potentiometric sensor, namely, Pt|Hg|Hg2 FUM|graphite, where FUM represents the fumarate
Received 5 May 2011 ion was developed for the measurement of fumaric acid in foods. The characteristics, performance
Received in revised form 21 January 2012 and application of the technique are described. The sensitivity of the electrode to FUM was
Accepted 19 February 2012
(29.2 ± 0.6) mV decade1 over a wide concentration range (7.5  107–1.0  102 mol L1), at pHs
Available online 27 February 2012
between 6.0 and 9.0, and the detection limit was 5.8  107 mol L1. The electrode is easily constructed
at a relatively low cost, has a fast response time (within 15–30 s) and can be used for up to 5 months
Keywords:
without showing any significant changes in performance. The sensor displayed good selectivity in the
Fumarate-sensitive electrode
Potentiometry
presence of a variety of other anions (carboxylates and inorganic anions). It was successfully used for
Gelatin dessert powders the determination of fumaric acid in powdered food products including gelatin dessert powders, instant
Instant pudding powder pudding powder and ice cream powders.
Ice cream powders Ó 2012 Elsevier Ltd. All rights reserved.

1. Introduction
Food additives have been used for centuries. Food preservation
began when man first learned how to store food between harvests
and with the salting and smoking of meat and fish. The Egyptians
used colourings and flavourings, and the Romans used saltpeter
(potassium nitrate), spices and colourings for preservation and to
improve the appearance of foods. Cooks have regularly used baking
powder as a raising agent, thickeners for sauces and gravies, and
colourings, such as cochineal, to transform raw materials into
foods products that are safe, wholesome and enjoyable to eat.
The underlying principles of modern food manufacturing methods
remain much the same as those used in traditional home cooking
for food preparation and preservation (Saltmarsh, 2000).
Over the last 50 years, developments in food science and tech-
nology have led to the discovery of many new substances that
can fulfill numerous functions in foods. Widely used food additives
include emulsifiers in margarine, sweeteners in low-calorie prod-
ucts and a wide range of preservatives and antioxidants that inhi-
bit spoiling and rancidity whilst retaining flavour.
Fumaric acid is a food additive (acidulant) that has been widely
used since 1946 because it is inexpensive and non-toxic (Singhal,
Kulkarni, & Rege, 1997). In addition, fumaric acid is an important
indicator of microbial spoilage as well as of the authenticity of
juices (Evans, VanSoestbergen, & Ristow, 1983). The major food
groups contributing to the dietary intake of fumaric acid and its
⇑ Corresponding author. Tel.: +55 16 33016665; fax: +55 16 33227932.
E-mail address: pezza@iq.unesp.br (L. Pezza).
salts (sodium, potassium and calcium salts) are desserts and sugar
confectionary. The maximum permitted level in dry powdered des-
sert mixes, gel-like desserts and fruit flavoured is 4000 mg kg1.
The acceptable daily intake (ADI) values for fumaric acid and
fumarate salts, determined by the Joint FAO/WHO Expert Commit-
tee on Food Additives (JECFA), is 60 mg kg1.
Various analytical methods for the determination of fumaric
acid in food products have been reported in the literature. These
include polarography (AOAC Official Method 968.16., 2000), high
performance liquid chromatography (DaPorto & Munari, 1989;
DolencSturm, Stampar, & Usenik, 1999; Durst, Wrolstad, & Krue-
ger, 1995; Evans et al., 1983; Lee, 1993; Masson, 2000; Montanari,
Perretti, Natella, Guidi, & Fantozzi, 1999; Trifirò et al., 1997), gas
chromatography (Tsuda, Nakanishi, Morita, & Takebayashi, 1985),
gas chromatography–mass spectrometry (Ayaz, Kucukislamoglu,
& Reunanen, 2000; Keou-Ng, Lafontaine, & Harnois, 2000), capillary
isotachophoresis (Blatńy, Kvasnička, & Kenndler, 1996) and capil-
lary isotachophoresis-capillary zone electrophoresis (Kvasnicka &
Voldrich, 2000). However, many of these methods are often com-
plicated and time consuming or require expensive equipment.
There is therefore a demand for alternative simple, low-cost, sensi-
tive and rapid methods for the determination of fumaric acid in
food products.
Potentiometric detection using ion-selective electrodes (ISEs)
has been shown to be effective for the analysis of pharmaceutical
formulations, biological materials and food samples. The advanta-
ges of these sensors include speed, simple preparation procedures,
applicability to coloured and turbid solutions, relatively short re-
sponse times, reasonable selectivity, wide linear dynamic range,

0308-8146/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
doi:10.1016/j.foodchem.2012.02.104
484 A.O. Santini et al. / Food Chemistry 134 (2012) 483–487
low cost and ability to be interfaced within automated systems
(Lewenstam, Maj-Zurawska, & Hulanicki, 1991).
To the best of our knowledge, there have been no previous re-
ports concerning either the use of potentiometric sensors for the
determination of fumaric acid in food products, or techniques for
the measurement of fumaric acid in dessert powders.
The present work describes the development, evaluation and
application of a simple and low-cost potentiometric fumarate ion
sensor immobilized in a graphite matrix (Pt|Hg|Hg2 FUM|graphite,
where FUM represents the fumarate ion), for the determination of
fumaric acid in powder matrices including gelatin dessert, instant
pudding and ice cream powders. Advantages of the proposed sen-
sor include simplicity, fast response, reasonable stability and
repeatability and very low detection limit.
2. Experimental
2.1. Reagents
High purity deionized water (resistivity = 18.2 MX cm) ob-
tained from a Milli-Q Plus system (Millipore Corp., Bedford, MA,
USA) was used throughout. All reagents employed were analytical
grade and obtained from E. Merck (Darmstadt, Germany), except
fumaric acid and fumaric acid disodium salt (sodium fumarate),
which were supplied by Sigma (St. Louis, MO, USA).
The sodium fumarate (Na2FUM) stock solution (2.0  101
mol L1) was prepared in deionized water at constant pH
(6.0 ± 0.1). This solution was stable for at least 3 months when
stored at 5 °C.
Standardizations of carbonate – free sodium hydroxide, nitric
acid and sodium nitrate solutions were performed as described else-
where (Moraes et al., 1996). Metallic mercury was purified accord-
ing to a previously reported procedure (Moraes et al., 1996).
Hg2FUM was prepared in aqueous solution by mixing the corre-
sponding nitrate with an excess of Na2FUM. The resulting precipi-
tate was filtered through a sintered glass funnel, washed with
deionized water until it was free of nitrate, and then dried to con-
stant mass over calcium chloride in a vacuum desiccator at room
temperature.
2.2. Electrode preparation and conditioning
The Hg2 FUM electrode was prepared as follows: Hg2 FUM (1.9 g)
and metallic mercury (ca. 0.3 g) were mixed in an agate mortar and
the material was crushed until a homogeneous solid was obtained.
Pure powdered graphite (1–2 lm particle size, Aldrich, USA) was
then added in a mass ratio of 50% and the crushing process was con-
tinued until complete homogenisation was achieved. A portion of
the resulting solid was transferred to a press mould, and compressed
at 9 tons for about 6 min. The resultant black pellet (1.6 mm thick,
12 mm o.d., and 0.8 g mass) was affixed to the tip of a glass tube
(external diameter 12 mm and length 8 cm) using silicone-rubber
glue (‘‘Rhodiastic’’, Rhône-Poulenc, France) and allowed to dry for
48 h. Sufficient metallic mercury (ca. 0.6 g) was then introduced into
the tube to produce a small pool on the inner pellet surface; electrical
contact was established through a platinum wire plunged into the
mercury pool and a subsequent conductor cable. The electrode
was then sealed using the procedure previously reported by the
authors (Pezza et al., 2001).
Elemental mercury and its compounds are potentially toxic (all
forms of mercury can enter the body) and as such are considered
to be hazardous wastes. All operations involving the use and manip-
ulation of metallic mercury and mercury (I) compounds were there-
fore conducted in a laboratory hood that had been checked for
adequate face velocity and proper operation. During the handling
of metallic mercury and mercury (I) compounds the operator used
safety glasses, appropriate protective clothing and latex or nitrile
gloves. Disposal of waste mercury employed appropriately marked
containers that were periodically by a specialist disposal company.
These safety precautions avoided (or at least minimised) the
operator’s exposure to mercury. In use, the proposed sensor does
not present any significant risk to health, and can be considered safe.
When not in use, the electrode’s pellet was kept immersed in a
small volume of freshly prepared 0.010 mol L1 Na2FUM solution at
pH = 6.0 ± 0.1 whose ionic strength (l) was adjusted to 0.500
mol L1 with NaNO3.
Prior to each experiment, the external surface of the pellet was
washed with deionized water and dried with absorbent paper.
2.3. Instruments
The electromotive force (emf) values were read to the nearest
0.1 mV with a Metrohm model 692 pH/ion metre (Metrohm Ltd.,
Herisau, Switzerland).
The reference electrode was a Metrohm Ag|AgCl double junction,
model 6.0726.100. The pH of aqueous solutions was adjusted and mon-
itored with the aid of a Metrohm pH electrode, model 6.0234.100. A
thermostated titration cell (25.0 ± 0.1 °C) was employed.
Volume measurements (± 0.001 mL) were performed using a
Metrohm model 665 automatic burette.
All experiments were performed at 25 ± 1 °C in a temperature-
controlled room.
2.4. Potentiometric cell
The following cell was used,
The value of x was in the range 1.0  107–1.0  101 mol L1.
The ionic strength (l) of the cell compartments was kept constant
at 0.500 mol L1, and the outer compartment of the reference elec-
trode was periodically refilled with fresh NaNO3 solution.
The performance of the Hg2 FUM electrode was assessed by
measuring the emf of the aforementioned cell for Na2FUM solution
concentrations in the range 1.0  107–1.0  101 mol L1 Na2FUM
solutions. These solutions were freshly prepared by serial dilution
of a 2.0  101 mol L1 stock standard solution with deionized
water, at constant pH (6.0 ± 0.1) and with l adjusted to
0.500 mol L1 with NaNO3. The emf readings were observed while
stirring the solutions, and recorded when they became stable. A
typical electrode calibration plot is shown in Fig. 1.
2.5. Determination of fumaric acid in powdered food samples
Powdered food samples, claimed to contain fumaric acid as an
additive at permitted concentrations, included gelatin dessert, in-
stant pudding and ice cream. The powders were purchased in local
markets and all were analysed prior to the state expiry date.
2.5.1. Determination of fumaric acid in gelatin dessert powders,
instant pudding powder and ice creams powders
An appropriate amount (typically between 0.5 and 0.6 g) of a
certain powdered food sample was weighed out, dissolved in
 A.O. Santini et al. / Food Chemistry 134 (2012) 483–487 485

5.8  107 mol L1. The sensor response displayed good stability
and repeatability over the period of the tests, as demonstrated by
the standard deviation values obtained (Table 1).

3.2. Response time and lifetime of the electrode

The response time and the lifetime of a sensor are of critical

Fig. 1. Calibration graph for the proposed fumarate -sensitive electrode (pH = 6.0;
l = 0.500 mol L1 adjusted with NaNO3; T = 25 °C).
80 mL of deionized water, homogenised for 3 min in an ultrasonic
bath and transferred to a 100 mL volumetric flask. The ionic
strength of the samples was adjusted to 0.500 mol L1 with NaNO3
and the pH to 6.0 ± 0.1 with 102 mol L1 HNO3 or 102 mol L1
NaOH before volume completion using deionized water. A 10 ml
aliquot was employed for analysis with the fumarate-sensitive
electrode using the standard additions method (with multiple
additions).
3. Results and discussion
3.1. Electrode response
The experiments carried out as described above (Section 2.4)
resulted in the following linear relationship between the measured
emf (E, in mV) and the FUM ion concentration:
E ¼ E0  S log ½FUM
Where E0 is the formal cell potential and S represents the theo-
retical Nernst slope (29.58 mV/decade, for divalent anions, at
25 °C). Potentiometric parameters and other features of the Hg2
FUM electrode are provided in Table 1. The above calibration equa-
tion and the slope value (Table 1) show that the electrode has a
near-Nernstian response to the fumarate ion in the range
7.5  107–1.0  102 mol L1. The limit of detection, determined
from the intersection of the two extrapolated segments of the cal-
ibration graph (Fig. 1) (Buck & Lindner, 1994), was
importance for analytical applications. According to IUPAC recom-
mendations, the response time may be defined as the time taken
the addition of analyte to the sample solution and the attainment
of a limiting potential (Buck & Lindner, 1994).
The response time of the electrode was determined by measur-
ing the time required to achieve a steady state potential (with-
in ± 0.2 mV/min), 5.0  107–1.0  102 mol L1 Na2FUM
solutions at pH 6.0 ± 0.1. Stable potentials were obtained within
15–20 s at high concentrations (P1.0  103 mol L1) and after
about 30 s at concentrations near the detection limit. The experi-
mental results showed that the lifetime of the electrode was
around 5 months, during which a total of 580 determinations were
performed. The sensor was used daily throughout the period, and
when not in use was stored in freshly prepared 0.010 mol L1 Na2-
FUM solution (pH = 6.0 ± 0.1 and l = 0.500 mol L1, adjusted with
NaNO3). No significant changes in the working concentration
range, slope and response time were observed during this period.
3.3. Influence of pH
The dependence of the electrode potential on pH was tested
over the pH range 4.0–10.0 for FUM ion concentrations of
1.0  103 and 1.0  104 mol L1. The pH was adjusted using di-
luted solutions of HNO3 or NaOH. The potential response remained
almost constant over the pH range 6.0–9.0 (Fig. 2) which was
therefore taken to be the working pH range of the electrode.
At pH values below 5.0, significant fractions of the fumarate ion
(pKa1 = 3.02 and pKa2 = 4.38) (Haynes, Jones & Motherwell)
changes to the corresponding monoprotonated and diprotonated
forms, which are not detected by the electrode. For pH > 9.0, the
hydroxide ion interferes with the electrode’s response. In high
pH media, probably OH competes with fumarate in the electrode
process and alters the potentiometric response of the proposed
sensor.

Table 1
Potentiometric response characteristics of the fumarate-sensitive sensor. Conditions:
T = 25.0 ± 0.1 °C; pH = 6.0 ± 0.1; l = 0.500 mol L1 (NaNO3).
a
Slope (mV decade1) (29.2 ± 0.6)
0 a
Intercept, E (mV) 40.1 ± 1.2
Linear range (mol L1) 7.5  107 – 1.0  102
Detection limit (mol L1) 5.8  107
Response time (s) 15–30
Working pH range 6.0–9.0
Analytical application Fumaric acid determination in food products
a
Average value ± SD of 46 determinations over a period of 5 months. Number of Fig. 2. Influence of pH on the response of the electrode: (j) 1.0  103 mol L1
data points: 20–25. Mean linear correlation coefficient: 0.998 ± 0.006. FUM, (d) 1.0  104 mol L1 FUM; l = 0.500 mol L1 adjusted with NaNO3; T = 25 °C.
486 A.O. Santini et al. / Food Chemistry 134 (2012) 483–487

Table 2 tion of fumaric acid in these matrices without any need for prior
Selectivity coefficients (KFUM,M) for different anions, determined extraction procedures.
using the matched potential method (see Section 3.4. for details).

Anion KFUM,M
3.5. Robustness of the technique
Formate 7.2  104
Acetate 8.1  104
The robustness of the method was investigated by the evaluat-
Propionate 7.6  104
Citrate 5.8  104
ing the influence of small variations of some of the most important
Lactate 8.8  104 procedure variables, including pH, potential range and measure-
Folate 1.8  103 ment time. Preliminary inspection of these results obtained under
Benzoate 2.3  103 different conditions indicated that the technique was fairly robust,
Sorbate 1,2  103
although the pH of the measuring solution should be maintained in
Salicylate 3.1  103
Dehydroascorbate No interference the range 6.0–9.0.
Chloride 3.6  102
Sulphate 3.1  105
3.6. Analytical application
Borate 4.3  105
Perchlorate No interference
Nitrate No interference In order to check the usefulness of the proposed sensor for mea-
surements involving real samples, fumaric acid was analysed in
foods (powdered soft drinks, gelatin dessert powders, instant pud-
3.4. Electrode selectivity ding powder and ice cream powders) using the standard additions
method (with multiple additions). The methodology was validated
The potentiometric selectivity coefficient, which reflects the rel- using five different powdered food matrices (two gelatin dessert
ative response of a sensor to the primary ion compared to other powders, one instant pudding powder and two ice cream pow-
ions, present in the solution, is perhaps the most important charac- ders). Precision was determined by performing analyses in tripli-
teristic of any ion-sensitive sensor. The potentiometric selectivity cate for each type of sample.
coefficients for the fumarate-sensitive electrode (KFUM,M) were Accuracy is a main requirement of any analytical technique and
determined, for a number of interfering ions, using the matched can be assessed in several ways (Prichard & Barwick, 2007): use of
potential method (MPM) (Gadzekpo & Christian, 1984). In this pro- an appropriate certified reference material; recovery of material
cedure, the selectivity coefficient is defined by the ratio of the spiked into a blank matrix; standard additions of the compound;
activity of the primary ion relative to an interfering ion, when they comparison with results obtained using another validated or refer-
generate identical potentials in the same reference solution. In the ence method. In the present case, the target analyte was not pres-
MPM, both monovalent and divalent ions are treated in the same ent in simple matrices and no blank media are available.
manner and the valence of the ions does not influence the selectiv- Furthermore, a thorough literature survey revealed an absence of
ity coefficient. Furthermore, the MPM can be used irrespective of analytical methods for the determination of fumaric acid in dessert
whether the electrode slopes are Nernstian or linear (Bakker, Bühl- powders (gelatin, pudding, ice-cream, etc.). The accuracy of the
mann, & Pretsch, 1997). method was therefore established using the standard additions
The MPM-selectivity coefficients (KFUM,M) were determined technique, spiking pre-analysed samples (in triplicate) with fuma-
using an: Initial reference solution (pH = 6.0 ± 0.1) containing ric acid at two different concentration levels.
0.500 mol L1 NaNO3 as the supporting electrolyte and 1.0  Table 3 shows the precision and accuracy obtained for measure-
105 mol L1 of the primary ion (fumarate). The selectivity coeffi- ments (in triplicate) of fumaric acid in five types of powdered
cients were calculated from the concentration of the interfering
ion (M) that, induced the same potential change (Demf = 15.0 mV)
as that induced by increasing the concentration of the primary ion. Table 3
The values of KFUM,M obtained (Table 2) showed that the Hg2 FUM MMethod validation data (precision and accuracy) for fumaric acid in five types of
electrode presented good selectivity considering all of the organic powdered food.

acid anions tested. Benzoate and sorbate are also used as food pre- Sample Measured Spike Recovery (%, RSD
a b
servatives, while citrate is found in many food products. No inter- concentration (g kg1) ±SD) (n = 3) (%)
ference was noted for most of the components commonly found (g kg1)

together with fumaric acid in the samples matrices investigated Gelatin dessert 1.89 ± 0.05 2 97.4 ± 5.8 5.9
(powdered food samples: gelatin desserts, instant puddings and powder 1
(Cherry)
ice creams) such as sodium saccharin, sodium cyclamate, aspar-
4 101.7 ± 3.2 3.1
tame, acesulfame K, lactose, fructose, glucose, sucrose, maltodex- Gelatin dessert 1.93 ± 0.06 2 99.3 ± 2.8 2.8
trin, citric acid, sodium benzoate, potassium sorbate, sodium powder 2
citrate, gum arabic, ammonium sulphate of caramel, Sunset Yel- (Raspberry)
4 98.1 ± 4.9 5.0
low, Yellow 5, Ponceu 4R, FD&C Red 40, FD&C Blue 1, Caramel Col-
Instant pudding 1.67 ± 0.04 2 100.5 ± 2.2 2.2
our, sodium phosphate, tricalcium phosphate, silicon dioxide and powder 1
titanium dioxide. (Chocolate)
Sulphate and borate have very low selectivity coefficients (Table 4 101.1 ± 3.6 3.6
2), while no interference at all is caused by either nitrate or per- Ice cream powder 1 1.82 ± 0.03 2 100.3 ± 1.9 1.9
(Strawberry)
chlorate, which can therefore be used as background electrolytes
4 99.2 ± 2.4 2.4
or ionic strength adjusters for fumarate solutions before perform- Ice cream powder 2 1.74 ± 0.05 2 102.3 ± 5.9 5.8
ing potentiometric measurements. (Chocolate)
Some interference by chloride ion might be expected. How- 4 101.7 ± 4.6 4.5
ever, the chloride ion is seldom present at significant concentra- a
Average of three independent analyses (n = 3) ± the corresponding standard
tions in the food samples used in this work, so that the deviation (SD). Expressed as fumaric acid.
b
proposed sensor can therefore be used for the direct determina- Relative standard deviation (RSD).
 A.O. Santini et al. / Food Chemistry 134 (2012) 483–487
foods. The presence of fumaric acid in each food was stated on the
labels, but the concentrations were not provided. The results ob-
tained showed that in all cases the levels of fumaric acid were be-
low those permitted by Brazilian legislation (Brasil., 1999).
Average recoveries were in the range 97.4–102.3% range (RSDs
of 1.9–5.9%, n = 3), showing the absence of any significant matrix
effects and the efficacy of the method.
4. Conclusions
The results obtained demonstrate the suitability of a new
potentiometric method employing a fumarate-sensitive electrode
immobilized in a graphite matrix for the quality control of fumaric
acid in foods.
The proposed potentiometric sensor is relatively easy to pre-
pare, and exhibits a long lifetime, high sensitivity and a wide dy-
namic range. Good selectivity, low detection limit, rapid response
and low cost make this electrode suitable for the analysis of fuma-
ric acid in food products including gelatin dessert powders, instant
pudding powders and ice cream powders.
Acknowledgments
We would like to thank the Brazilian agencies FAPESP, CNPq,
CAPES and FUNDUNESP (Brazil), for financial support.
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