Determination of Fe, Cu, Zn, Mn, and Pb
in D.O.C. M ntrida wines
M. Jos~ Gonzfilez, M. Carmen Martinez Para, and M. Victorina Aguilar
Departamento Farmacia, Nutrici6n y Bromatologia, Facultad de Farmacia, Uuiversidad de Alcalfi de Henares,
Apto. 20, Alcalfi de Henares, Madrid, Spain
Bestimmung von Fe, Cu, Zn, Mn, und Pb
in M~ntrida-D.O.C.-Weinen
Zusammenfassung. Es wurden 14 Weine aus Mtntri-
da, Toledo (Spanien) mit Hilfe der Atomspektropho-
tometrie (AAS) auf ihre Konzentration an Fe, Cu, Zn
und Pb untersucht und zwar einesteils die unbehandel-
ten Proben direkt und andernteils die durch feuchten
AufschluB erhaltenen Proben. Es sollte daraus ge-
schlossen werden, ob organische oder anorganische
Matrixeffekte bestehen. Weiterhin wurde der EinfluB
der Weinerzeugung auf den Gehalt an Spurenelemen-
ten sowie die Stabilitfit und Zutr~iglichkeit der Weine
in bezug auf den Gehalt an toxischen Metallen ver-
folgt.
Summary. Fourteen wines from Mtntrida (Toledo,
Spain) were examined by atomic absorption spectro-
photometry (AAS) in order to measure the concentra-
tions of Fe, Cu, Zn, Mn, and Pb, using two different
methods of sample preparation for the metal analyses:
analysis of untreated samples and analysis of wet-
mineralized samples. This was camed out in order to
elaborate a suitable method for their determination
after studying the possible matrix effects originating in
either the organic or inorganic components. The influ-
ence of the vinification system on trace metal content
and the stability and salubrity of these wines in rela-
tion to certain toxic metal concentrations was also ex-
amined.
Introduction
When defining the organoleptic and enological prop-
erties of any wine, different organic and inorganic
components come play a role. The quantification of
these components is very important, not only analyti-
cally and technologically, but also with regard to their
Offprint requests to: M.J. Gonz~.lez
toxic and nutritional effects. Mineral elements are nat-
ural constituents of wine and directly affect the final
quality and characteristics of the wine. The enological
effects of some metallic elements affect wine stability
and commercial acceptability, for example iron and
copper, which at certain levels cause so-called ferric
and cupric cloudiness. Others, like manganese, are, ac-
cording to some authors [1] indicative of the geo-
graphic wine-producing region. Therefore by measur-
ing these elements, it is possible to characterize a con-
trolled wine-producing region. The evaluation of zinc,
and, especially lead, which is cumulative, is particu-
larly important.
A great number of mineral elements present in
wine at the last stage of its elaboration depend on dif-
ferent factors, both natural and exogenous. Natural
factors include soil type and composition, grape
variety, climate, etc. Exogenous factors can derive
from the fermentation process, the vinification system
or from different kinds of contamination: including
fungicide use, corrosion of wine-making equipment,
and proximity to industrial plants, etc. [2, 3]. The ob-
ject of this study was to ascertain the amount of Fe,
Cu, Zn, Mn, and Pb present in Mtntrida wines and so
to establish a suitable method for their quantification
using AAS technique. Two sample preparation
methods were used: direct sample analysis without
pre-treatment and pre-preparation by wet mineraliza-
tion. Recovery test matrix interference studies for
both the simple wine sample and the mineralized
sample and, finally, a study of organic interferences
were performed to compare the two sample prepara-
tion methods.
Material and methods
To make this study representative of the Mtntrida region, wine
samples were taken from 14 registered wineries scattered throughout
the entire region. The samples were red and ros6 wines which had
been harvested in 1983 and 1984.
Metal element determinations were carried out with a model
2380 Perkin Elmer atomic absorption spectrophotometer equipped
with a model 561 register. Multi-element hollow cathode lamps were
used to determine Fe, Cu, Zn, and Mn and an electrodeless discharge
Z LebensmUnters Forsch (1988) 187:325 329
© Springer-Verlag1988
TaMe 1. Working conditions 3 Interference from ethanol
Ele- > Slit Flame Light Linear D L Sensiti- Pre- and other organic compounds
ment sett- type source work- vity cision
ing ing % CV Except for Cu and Mn determinations, wine presents
range
(nm) (nm) (Ixg/ml) (~tg/ml) (lxg/ml) matrix interferences for all of the elements studied. We
used the method of standard addition to examine the
Cu 324.8 0.7 AAFO HCL 05 0.008 0.040 2.0
Fe 248.3 0.2 AAFO HCL 05 0.016 0.190 3.1 effects produced by ethanol and other organic com-
Mn 279.5 0.2 AAFO HCL 03 0.007 0.087 2.6 pounds, such as tartaric acid, citric acid, sucrose and
Pb 283.3 0.7 AAFO EDL 20 0.030 0.700 1.8
Zn 213.9 0.7 AAFO HCL 01 0.002 0.010 4.5 glycerol, on AAS determination. These wine matrix
compounds were chosen because they are present in
A A F O air-acetylene oxidizing flame; H C L hollow cathode lamp;
E D L Electrodeless discharge lamp
wine and because they were also part of the composi-
tion of the synthetic wine solution used in this paper
for direct measurement. The results of this study show
lamp was used to determine Pb. The working conditions are shown that the presence of these compounds leads to differ-
in Table 1.
Element quantification was carried out with both sample types;
ent errors in quantity and signal production for the
the wet mineralized sample was determined after adding a sulphoni- majority of the elements (except Cu and Mn), falsifies
tric mixture and the untreated sample was determined after calibrat- readings and should, therefore be eliminated.
ing the apparatus with a synthetic wine. Pb and Mn were only deter-
mined in the mineralized sample, since their concentrations in the un- 4 Fe, Cu, Zn, Mn, and Pb content
treated sample were so low that interferences from other wine com-
ponents blocked any quantification. In order to examine the matrix
in D.O.C. Mbntrida wines
interferences and the influence of organic and inorganic compounds, Tables 2-6 show the results for both the direct wine
the method of standard addition was applied.
measurement and after sample pre-mineralization.
The results also indicate the presence of each element
Results and discussion in the red and ros6 wine during each harvest. To judge
1 Matrix interferences the significance of the results, a variance analysis was
performed. A parametric comparison test was used
After applying the method of standard addition, it for Pb and Mn and the other elements were studied
could be seen that none of these elements presented with a non-parametric comparison test.
matrix interference in the mineralized sample. Al-
though matrix interferences are obvious in the deter- Iron. This element is part of the wine mineral fraction
mination of Fe and Zn in untreated wine samples, they and, together with copper, is important, as much for
are not significant for Cu and Mn. Lead was not deter- its influence on wine conservation and stability as for
mined at this point, since the concentration at which its role in determining the organoleptic characteristics
it is found is so low that any interference will thwart of the wine [6]. Iron levels determined directly from
its quantification, as was proven experimentally. untreated samples vary a minor amount of between
1.84 and 10.64 rag/1 and between 2.37 and 10.21 in the
2 Recovery test; chemical interference mineralized samples. They are among the lowest re-
In order to corroborate the validity of the mineraliza- ported in the abundant literature on Fe in red and ros6
tion method, recovery test were carried out with both wines [1, 7, 8] and indicate a minor amount of contam-
the standard aqueous solution and the sample solu- ination during the wine-making process (Table 2).
tion. The results for Cu and Pb were optimal. How- Since the Fe concentration present in the wines
ever, Fe, Mn and Zn were too low (below 90%), prob- does not exceed 10 mg/1, there should be no danger of
ably because of the influence of other metallic ions on ferric cloudiness [9] and the stability of these wines
their determination. Some authors [4, 5] indicate the should be assured.
existence of interferences due to Pb, Cu, Na, K, and The acidity in ros6 wines is higher than in red
Fe. Once this hypothesis had been checked, we were wines, since sulphur dioxide is also higher and conse-
able to confirm that the presence of these elements cuently ros6 is more corrosive than red wine. For this
negatively affected the determination of the samples reason, Fe concentrations in ros6 wines are higher,
by lowering the signal produced at intervals between since the wine Fe content is enriched by corrosion of
0.5% and 8% (relative error). After this test, recovery the iron in the packaging, machinery, installations and
tests performed with standard aqueous solutions for utensils when the must and the wine come into contact
each isolated element with the object of eliminating [10].
possible interactions. The percentage recovery values
confirmed the above and were as follows: Mn, 95.5%; Copper. This metal is found in wine at concentrations
Fe, 97.5%, and Zn, 98%. varying between 0.1 and 0.5 mg/1 [11] although other

326
Table 2. Levels o f Fe in ros6 and red wines harvested in 1983 and authors have estimated copper levels to be closer to 1
1984 (mg/1)
or 3 mg/1 [12]. It is generally believed that the mean va-
Win- 1983 1984 lues taken between 0.2 and 0.3 mg/1, create no pro-
ery
Ros6 Rosb Red Red Rosb Ros+ Red Red blems in stability with respect to cupric cloudiness [6].
(m)' (d) u (m) (d) (m) (d) (m) (d) The legal limit for this element in Italian legislation is
1 rag/1 [13].
1 3.14 5.66 5.34 2.82 3.82 2.66 3.18 2.78
2 7.68 6.02 3.54 3.28 3.66 4.29 3.18 2.94 The differences in copper concentrations found
3 5.06 5.05 2.70 2.85 7.23 10.64 6.30 5.13 between the two sample methods used in this study
4 6.50 3.23 3.53 2.91 7.22 6.01 4.66 4.76
5 4.29 4.18 2.78 2.37 2.37 4.50 5.12 4.50 were not statistically significant. Directly analysed un-
6 2.64 2.48 3,69 3.19 8.20 5.15 10.21 9.41 treated samples give a maximum copper concentrati-
7 3.65 3.23 4.50 3.12 6.10 3.86 4.56 4.44
8 2.80 2.74 7.12 3.26 6.74 4.50 4.47 1.84 on of 5.56 mg/1 as compared with values as great as
9 10.00 7.73 2,92 2.84 6.91 4.29 4.53 4.24
10 3.82 3.66 4.05 3.61 7.39 4.08 8.95 4.94
6.21 mg/1 detected in the mineralized samples. Only
11 3.43 3.24 9.34 6.01 6.44 6.10 one sample showed a value as high as 6.21 rag/1 and
12 4.37 2.67 2.58 2.43 6.26 4.29 6.28 3.26
13 8.74 5.99 2.38 2.03 4.80 3.71 4.50 3.77 two samples exceeded 1 rag/1. These exceptional quan-
14 4.56 3.93 4.48 3.21 3.70 2.30 tities are probably due to accidental contamination of
" Mineralized wine measurement
the wine or to prolonged contact with copper equip-
b Direct wine measurement ment [1]. Since most of the samples did not exceed
0.3 rag/l, 85% showed no cupric cloudiness. These va-
lues are similar to those reported in the literature [1,
14, 15]. Only three samples exceeded the Italian legal
Table 3. Levels o f Cu in ros6 and red wines harvested in 1983 and threshold of 1 mg/1. As with iron, copper levels, in ros6
1 9 8 4 (mg/1) wines were higher than in red wines for identical rea-
Win- 1983 1984
sons (Table 3).
ery
Ros6 Ros6 Red Red Ros6 Ros6 Red Red Zinc. Only traces of this element were found in these
(m) (d) (m) (d) (In) (d) (m) (d)
wine. The levels of zinc found in the untreated wine
1 0.103 0.100 0.010 0.050 1.070 0.750 0.170 0.510
samples varies between 0.16 and 2.55 mg/1 and bet-
2 0.105 0,110 0,050 0,042 0,015 n.d. 0.050 n.d. ween 0.18 and 2.73 mg/1 in the mineralized samples.
3 0.052 n.d. 0.012 0.010 0,150 0,160 0,290 0.046
4 n.d." n.d. 0.008 n.d. 0.210 0.020 0,050 n.d. These values fall within the range 0 . 1 5 4 rag/1 and are
5 0,055 0.036 0.015 0,017 0.230 0,250 0,021 0.029 considered to be normal [16]. However, they are lower
6 0,065 0,065 0,020 0,025 0.630 0.670 0,250 0,290
7 0,040 0,048 n.d. n.d. 0.130 0,120 0,130 0.100 than zinc concentrations mentioned in other studies
8 0.148 0,130 0,130 0,160 0.500 0.310 0,350 0,360 [17-19]. The legal limit for zinc in Italy is 5 mg/1, and
9 0.232 0.220 0,020 0.040 0,190 0.170 0,510 0,270
10 0.007 n.d. 0.010 0.020 0.035 0,060 0,190 0.460 this level is not exceeded by any of the M6ntrida wines.
11 - - 0.430 0,230 1,600 1.820 0.230 0,270
12 0,084 0.054 n.d. n.d. 5.560 6,210 0,490 0,490
High concentration of this element suggest that it is
13 n.d. n.d. n.d. n.d. 0.110 n.d. 0,050 n.d. due to anti-parasitic treatment residues or zinc sul-
14 0,280 0.300 0,053 0.050 - n.d. n.d.
phate used together with potassiumferrocyanide, al-
" n . d . , levels below the detection limit though establishing the real cause it difficult [14]
(Table 4).
The differences between the values for the two dif-
ferent sample techniques were statistically significant.
Table 4. Levels o f Z n in ros6 and red wines harvested in 1983 and The lower levels found in the untreated wine sample
1984 (mg/dl)
can be explained by wine matrix interferences, in
Win- 1983 1984 which organic compounds provoke negative errors
cry
Ros6 Ros~ Red Red Ros6 Ros~ Red Red and ethanol provokes positive errors. Zinc concentra-
(m) (d) (m) (d) (m) (d) (m) (d) tions are statistically similar in both red and ros6
wines.
1 0.84 0.60 0.81 0,80 0.52 0.48 1,07 0.61
2
3
0.38
0.87
0.33
0.46
1.00
0.58
0.55
0.56
0.38
0.45
0.31
0.36
0.54
4.23
0.46
0.35
Manganese. Manganese levels in wine are generally
4 0.55 0.53 0.58 0.52 0.38 0.30 0.52 0.25 low, and some authors believe they are characteristic
5 0.48 0.53 1.00 0.98 0.35 0.29 0,34 0.29
6 0.59 0,62 0.77 0.88 0.49 0.39 0.98 0.53
of a specific wine-growing region, since the amount of
7 0.63 0.49 0.72 0.64 0.43 0.37 1.16 0.74 Mn in the soil is reflected in the final wine content [1].
8 1.99 0.96 0.98 0.79 0.65 0.58 1.21 0.65
9 0.61 0.48 0.98 0.87 0.76 0.69 0.18 0.64
Manganese concentrations in these wines varied bet-
10 0.49 0.34 0.42 0.36 0.18 0.16 0.50 0.26 ween 0.9 and 2.85 mg/1, which is similar to most of the
11 0.75 0.87 1.94 1.58 1.45 0.78
12 2.73 2.52 0.65 0.64 0.53 0,46 1.44 0.41 values obtained by other authors [1, 19-21]. However,
13 0.51 0.47 0.54 0.51 0.51 0.41 0.61 0.44 we are not in agreement with Interesse et al. [1], since
14 0.55 0.51 0.77 0.65 - 0.55 0.34
all the values are similar (Table 5).

327
T a b l e 5. Levels o f M n in ros6 a n d red wines h a r v e s t e d in 1983 a n d its the lead content in wine to 0.3 mg/1, a level that
1984 (mg/1)
was surpassed by three of our samples. However,
Winery 1983 1984 L'Office International de la Vigne et du Vin tolerates
Ros6 Red Ros6 Red levels of 0.6 mg/1 and these wines are within the nor-
mal and safe values (Table 6). The differences between
1 0.90 1.64 0.99 2.44
2 1.80 2.43 1.21 2.40 Pb levels in red and ros6 wines were not statistically
3 1.27 2.09 1.57 2.82 significant, although a slightly higher content can be
4 1.29 1.11 1.05 0.97
5 1.26 2.06 1.28 1.79 established in ros6 wines. This is a result of the ros6
6 1.42 2.17 1.60 1.70
7 1.28 1.15 1.37 2.40
vinification process when the grape juice is fermented
8 1.60 1.98 1.65 2.00 without the skins (which contain lead attracting
9 1.76 2.07 1.48 2.16
10 1.30 1.92 1.49 2.57 yeast). Therefore, during the making of red wine a
11 2.50 1.77 2.85 greater quantity of this element is removed when the
12 t.41 2.34 1.27 2.38
13 1.24 2.01 1.24 2.02 grape-skin and, obviously, the lead-attracting yeasts
14 1.11 1.95 2.32 are finally extracted; this is not done when making
ros6 wine and the Pb remains in solution [23].

T a b l e 6. Levels o f P b in ros6 a n d red wines h a r v e s t e d in 1983 a n d

1984 Conclusions
Winery 1983 1984 The results of this study allow the following conclu-
Ros6 Red Ros+ Red sions to be drawn:
1 n.d. n.d. n.d. n,d. 1. The direct method for analysing wine samples is
2 0.17 0.10 0.17 0.31 only useful in the determination of those elements
3 n.d. 0.03 0.20 0.27
4 n.d. n.d. 0.21 0.19 for which wine presents no matrix interferences,
5 n.d. n.d. 0.22 0.15 such as Cu and Mn; consequently the method is
6 n.d. 0.11 0.19 0.12
7 0.11 0.21 0.18 0.12 valid only when a quick and superficial study is
8 0.34 n.d. 0.21 0.08
9 0.01 0.04 0.013 0.10
performed. Therefore, wines need to undergo pre-
t0 n.d. 0.09 0.21 0.25 vious pre-treatment and mineralization before the
11 n.d. 0.24 0.08
12 0.09 n.d. 0.15 0.12 mineral element determination is carried out in
13 0.04 n.d. 0.52 0.22 order to remove any organic matter present in the
14 n.d. n.d. 0.11
wines.
n.d., levels u n d e r detection limit 2. Sample mineralization offers the advantages of
higher accuracy and security in comparison with
direct sample measurements, in all cases, even
Manganese content in wines depends on the type though the latter is faster. We believe this type of
of vinification. It is higher in red wines because during digestion is appropriate for the elimination of ma-
the wine-making process the must remains in contact trix interferences in AAS determination of these
for a longer period of time with the solid parts of the mineral elements. Chemical interferences cannot be
grapes, which have a higher Mn content and so dissol- eliminated by mineralization, so specific separation
ve the mineral salts present there [3]. techniques must be employed for each chemical.
3. The vinification process is a determining factor in
Lead. The lead content deserves special consideration the final wine mineral content. For red wine, Fe
among the toxic micro-elements that are presented and Cu levels are higher than those of ros6, while
being studied because of its cumulative character [13]. the Mn content in ros6 wine is higher than in red
The origin of this element in wines is basically exoge- wine.
nous, due to car exhaust fumes industrial explosions 4. The wines from the M6ntrida wine region are stable
and emissions, the use of organochloro anti-parasites with regard to Fe and Cu cloudiness.
and lead arsenates (which are now prohibited), the 5. Italian limits for Zn and Pb are not exceeded by any
proximity of a vineyard to ceramic factories, metallic samples from the M6ntrida region.
contamination from lead-containing tin capsules, etc.
[9, 22]. The normal lead content in wine varies bet-
ween 0.4 and 0.5 mg/1 [3, 10]. The wines studied in this References
paper have low lead levels. Lead concentration levels
1. Interesse FS, Lamparelli F, Alloggio VZ (1984) g Lebensm
were as high as 0.52 mg/1 in only one winery. The Unters Forsch 178:272-278
other wineries presented concentration levels that were 2. Cela R, Cabez6n ML, P6rez Bustamante JA (1983) An Quim
lower than 0.34 mg/1 for this element. Italian law lira- 79B:229-233

328
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Received December 11, 1987
Accepted March 7, 1988
329