Meat Science 86 (2010) 660–664

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Meat Science
j o u r n a l h o m e p a g e : w w w. e l s ev i e r. c o m / l o c a t e / m e a t s c i

Polycyclic aromatic hydrocarbons (PAHs) in two Spanish traditional smoked sausage

varieties: “Androlla” and “Botillo”
José M. Lorenzo ⁎, Laura Purriños, María C. García Fontán, Daniel Franco
Centro Tecnológico de la Carne de Galicia, Rúa Galicia No. 4, Parque Tecnológico de Galicia, San Cibrán das Viñas, 32900 Ourense, Spain

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

Article history: The polycyclic aromatic hydrocarbon (PAH) content in two traditional smoked sausages from Spain was
Received 29 September 2009 determined. Determination and quantification of PAHs in smoked sausages were performed by HPLC with
Received in revised form 15 December 2009 fluorescence detection. Results showed, that total mean levels of PAHs found were higher in “Androlla”
Accepted 14 May 2010
(36.45 µg/kg) than in “Botillo” (29.39 µg/kg) although no significant differences (P b 0.05) were observed. In
all examined samples content of phenanthrene was the highest in the two traditional sausage varieties. The
Keywords:
Polycyclic aromatic hydrocarbons
maximum level for benzo[a]pyrene (BaP) of 5 µg/kg in smoked meat products was not exceeded in any
Smoked meat products samples. BaP represented 1.3% and 1.2% in “Androlla” and “Botillo” samples, respectively of the total sum of
HPLC/FLD the 15 PAHs investigated in both sausages. Correlation statistic analysis (P b 0.01) showed that BaP was a
good marker for 6 IARC possible and probable carcinogenic PAHs in “Androlla” samples (RBaP/6IARC = 0.63)
and in “Botillo” samples (RBaP/6IARC = 0.96).
© 2010 The American Meat Science Association. Published by Elsevier Ltd. All rights reserved.

1. Introduction
“Androlla” and “Botillo” are a traditional raw-cured sausage
manufactured in Galicia (NW of Spain) and are included in the
“Catalogue of cured sausages and hams of Spain” (Anonymous, 1983).
Both sausages are very highly appreciated by consumers and are a
great success in the local markets. At the current, those sausages are
processed in a handmade or semi-industrial way, following absolutely
empiric traditional process (Lorenzo, Michinel, López, & Carballo,
2000). Traditional smoking of “Androlla” and “Botillo” sausages are
performed by incomplete firing wooden material, mainly from
Quercus robur, with a flame.
Smoking of meat and meat products is an ancient technology
which has been used for thousands of years. Nowadays, smoking
technology uses mainly the special effects of various sensory active
compounds contained in smoke for aromatisation of meat products
with a specific organoleptic profile, widely demanded on the market.
Incomplete wood combustion during process of smoking can produce
considerable amounts of polycyclic aromatic hydrocarbons (PAHs)
(Conde, Ayala, Afonso, & Gonzalez, 2005).
PAHs represent a group of organic compounds consisting of two or
more condensed aromatic rings that are widely geographically
distributed and remain in the environment for a long time (Howsam,
Jones, & Ineson, 2000). Decomposition phenomenon caused by UV
irradiation and by interaction with other present products causes the
⁎ Corresponding author. Tel.: +34 988 548 277; fax: +34 988 548 276.
E-mail address: jmlorenzo@ceteca.net (J.M. Lorenzo).
content of PAHs to diminish (Chen & Chen, 2005; Dennis, Cripps, Tricker,
Massey, & McWeeny, 1984; Simko, 1991). Nevertheless, the PAHs can
also penetrate into the smoked products, thus being protected from
light and oxygen, stabilizing their contents after a period of time (Simko,
& Knezo, 1992).
Potential health hazards associated with smoked foods may be
caused by carcinogenic components of wood smoke—mainly PHAs,
derivate of PAHs, such as nitro-PAHs or oxygenated PAHs, and to a
lesser extent also N-nitroso compounds and heterocyclic aromatic
amines. The health risk associated with dietary PAHs has been
evaluated by Scientific Committee on Food (SCF) (2002). In European
Commission (2005a) recommendations (EC 108/2005) the member
states have been advised to monitor 15 SCF priority PAHs together
with one additional PAH benzo[c]fluorene, recommended by the
JECFA (Join FAO/WHO Expert Committee on Food Additives, 2006).
At present, chromatographic techniques, mainly GC and HPLC, are
the dominant effective analytical tools in PAH determination. GC has
higher column efficiency and thus has an advantage for complex mixture
analysis, but HPLC can often have higher column selectivity, which is
more suitable for separation of isomeric compounds. Thus, both methods
should be viewed as complementary in the analysis of PAHs and they are
essential for precise and reliable analysis (Sinko, 2002, 2005).
Meat industry is interested in studying PAHs in smoked meat and
meat products, in order to improve food safety of products from
protected geographic areas and their compliance with the European
Union (EU) regulations. Smoked meat products represent a significant
part of the human diet in Galicia (NW of Spain). They are important
because of their good taste, high nutritional value, high level of
production and large variety of products.

0309-1740/$ – see front matter © 2010 The American Meat Science Association. Published by Elsevier Ltd. All rights reserved.
doi:10.1016/j.meatsci.2010.05.032
 J.M. Lorenzo et al. / Meat Science 86 (2010) 660–664
The aim of the present study was the investigation of the contents
of PAH compounds, which are recommended to be analysed by the
SCF and the European Food Safety Authority (EFSA), in two Spanish
traditional dry-cured sausage varieties: “Androlla” and “Botillo”.
2. Materials and methods
2.1. Sausage samples
For the study, fourteen and seventeen samples of “Androlla” and
“Botillo” sausage, made by 14 and 17 different industrial pork man-
ufacturers, respectively, were obtained in local markets.
For the manufacture of “Androlla”, low-quality pork (diaphragm,
inter-rib muscles, jowls, skin and, occasionally, lung) is minced and
salt, sweet and spicy paprika, garlic and sometimes onion are added.
The resulting mass is left standing for at least 24 h and then, it is
stuffed into pork rectum tripe in units of 20–25 cm length. After
stuffing, it undergoes a smoking-heating process for 8–10 days and
then a drying-ripening process for 1–2 months. “Botillo” is manufac-
tured from pieces of ribs, vertebrae and other pork bones with their
fleshy parts, skin and lard. The meat is mixed with salt, sweet and
spicy paprika, garlic and marjoram and is left to stand for 48 h and
then stuffed into pork caecum. It then undergoes a heating-smoking
process for 7–15 days, and afterwards it is left to dry and ripen for a
period of up to 3 months.
Once collected, the samples were transported to the laboratory
under refrigeration conditions (b4 °C). In order to prepare the
samples for analysis, after removing and discarding the outer casing
from pieces and eliminating the bone in “Botillo” samples, the edible
part was ground in a mincer machine (Moulinex/Swan Holdings Ltd.,
Birmingham, UK) until a homogeneous mass was obtained. The
samples were stored in air-tight bottles, frozen at −80 °C in dark, for
no longer than four weeks until analysis.
2.2. Reagents
The standard mixture NIST 1647d of 16 priority PAHs: naphthalene
(Naph), acenaphthene (Ace), acenaphthylene (Acy), fluorine (Fln),
phenanthrene (Phe), anthracene (Ant), fluoranthene (Flt), pyrene (Pyr),
benzo[a]anthracene (B[a]A), chrysene (Chr), benzo[b]fluoranthene
(B[b]F), benzo[k]fluoranthene (B[k]F), benzo[a]pyrene (B[a]P),
dibenzo[a,h]anthracene (DB[ah]A), benzo[g,h,i]perylene (B[ghi]P)
and indeno[1,2,3-cd]pyrene (I[cd]P) dissolved in acetonitrile was
supplied by National Institute of Standards and Technology (NIST,
Gaithesburg, MD, USA). Working standard solution (contents in the
range 0.01–60 ng/mL) was prepared in acetonitrile and stored at 4 °C.
Acetonitrile and n-hexane (HPLC grade) were purchased from Merck
Biosciences (Darmstadt, Germany) and dimethyl sulfoxide (DMSO)
was supplied from Panreac (Castellar del Vallès, Barcelona, Spain).
Before use, all glassware was washed with detergent, rinsed with
distilled water and acetone and then dried at 220 °C.
2.3. Extraction and clean-up
Homogenized meat samples were lyophilized (Labconco Corpo-
ration, Freezone, Kansas, USA). Then, 2 g (dry mass) of the lyophilate
was extracted three times with 25 mL for 1 h, 15 mL for 1 h and then
10 mL for 1 h with n-hexane in an ultrasonic bath (Branson Ultra-
sonics Corporation, 8510, Danbury, USA). The combined extracts
(50 mL) were centrifuged at 2500 rpm for 10 min, and the superna-
tant was decanted and concentrated to 5 mL under a nitrogen stream.
Then, the concentrated hexane extract was passed through a Waters
Sep-Pak C18 Silica Plus cartridge (particle size, 57.1 µm) and eluted
with 10 mL n-hexane at flow-rate of 1 mL/min. Then, the hexane
eluate (final volume, 15 mL) was extracted with 15 mL for 5 min,
10 mL for 5 min and then 5 mL for 5 min of DMSO that had previously
661
been equilibrated with n-hexane. The DMSO solution was diluted
with 75 mL water and passed through a Water Sep-Pak C18 Plus
cartridge (particle size, 90.7 µm) that had previously been activated
by passage of 5 mL of CH3CN followed by 10 mL of distilled water. The
eluates were discarded and the cartridge was eluted with 5 mL of n-
hexane. Prior to HPLC analysis, this hexane solution was concentrated
to dryness under a nitrogen stream in a vacuum evaporator (Zymark
Corporation, Turbovap LV, Massachusetts, USA), and the residue was
redissolved in 1 mL of CH3CN, which was then filtered through
0.45 µm pore-size Waters Acrodisc GHP filters (Waters Corporation,
Milford).
2.4. Separation and identification of the polycyclic aromatic
hydrocarbons
The separation and identification of the polycyclic aromatic
hydrocarbons were performed by HPLC methods in a Waters 2695
(Waters Corporation, Milford, MA, U.S.A.) with a Waters 2475
(Waters Corporation, Milford) programmable fluorescence detector
to identification and quantification of PAHs in meat sausages. The
column was a Thermo Hypersil Green PAH (100 mm × 4.6 mm i.d.,
3 µm) (Thermo Fisher Scientific, Bremen, Germany). The temperature
of the column during chromatographic separation was 35 ± 1 °C and
the setting for the programmable fluorescence detector was as follows
(excitation/emission wavelength): λ1 = 270/340 nm for 13.5 min
(naphthalene); λ2 = 270/340 nm for 2 min (acenaphthene, fluorene);
λ3 = 254/375 nm for 1.5 min (phenanthrene); λ4 = 260/420 nm for
2.5 min (anthracene, fluoranthene); λ5 = 254/390 nm for 6.5 min
(pyrene, benzo[a]anthracene, chrysene); λ6 = 260/420 nm for 6 min
(benzo[b]fluoranthene, benzo[k]fluoranthene, benzo[a]pyrene,
dibenzo[a,h]anthracene, benzo[g,h,i]perylene); λ7 = 293/498 nm for
2 min (indeno[1,2,3-cd]pyrene). The injection volume was 20 µL. The
chromatographic separation was carried out under the following
conditions: gradient elution (0 min—45% acetonitrile + 55% water;
26 min—80% acetonitrile + 20% water; 27 min—85% acetonitrile +
15% water; 28 min—90% acetonitrile + 10% water; 30 min—50%
acetonitrile + 50% water).The limits of quantitation (LOQ) for PAH
compounds were 0.007–0.041 µg/kg. All the samples and standards
were injected at least in duplicate. Repeatability tests were performed
by injecting a standard and sample consecutively six times in one day.
Reproducibility tests were also carried out by injecting the standard
and the sample twice a day for three days under the same ex-
perimental conditions. There were no significant differences (P b 0.05)
between the results obtained in either of these tests.
2.5. Statistical analysis
The experimental design was intended to determine the influence
of smoking process on PAH content in two Spanish traditional
sausages. The obtained results were statistically analysed using SPSS
15.0 statistical program package (Chicago, USA). One-way analysis of
variance (ANOVA) analysis was used to estimate the significance of
the differences between the means of total and individual PAH
content in samples analysed. To verify that BaP is a good marker for
other PAHs in analysed smoked meat products, the Pearson's cor-
relation coefficients (P b 0.05) between BaP values and the obtained
contents of analysed PAHs were calculated (SPSS 15.0).
3. Results and discussion
A total of 31 samples of meat product (17 “Botillo” and 14
“Androlla”) were analysed for contents of 16 priority PAHs (Tables 1
and 2). In some samples content was not detectable, i.e. concentra-
tion was below limit of quantitation (LOQ). Results showed that total
mean levels of PAHs found were higher in “Androlla” samples
(36.45 µg/kg) than in “Botillo” samples (29.39 µg/kg) although no
662 J.M. Lorenzo et al. / Meat Science 86 (2010) 660–664

Table 1
PAH contents (µg/kg dry weight) in “Botillo” samples.

Code Naph Ace Fln Phe Ant Flt Pyr B[a]A Chr B[b]F B[k]F B[a]P DB[ah]A B[ghi]P I[cd]P

B1 0.14 0.20 0.44 1.16 0.22 0.15 0.27 0.22 0.33 0.19 0.19 0.19 0.18 bLOQ b LOQ
B2 0.08 0.13 0.83 9.24 0.46 3.97 12.06 0.14 0.31 0.12 0.09 0.13 b LOQ 0.47 0.16
B3 0.06 0.09 0.09 0.67 1.71 1.45 45.34 0.26 0.29 0.42 0.12 0.25 b LOQ bLOQ 0.21
B4 0.06 0.08 0.08 0.20 0.21 0.93 10.00 0.13 0.21 0.49 0.08 0.24 0.78 3.40 0.23
B5 1.27 0.18 0.68 6.52 0.33 2.89 8.36 0.15 0.31 0.14 0.12 0.14 b LOQ bLOQ b LOQ
B6 14.90 0.41 2.81 13.89 2.43 2.00 5.53 1.06 1.13 b LOQ 0.50 0.43 b LOQ bLOQ b LOQ
B7 0.54 0.74 1.30 2.99 0.49 0.42 1.10 0.54 0.83 0.51 0.47 0.47 0.44 1.05 0.86
B8 0.08 0.12 0.10 0.27 0.75 1.14 33.15 0.30 0.33 0.53 0.12 0.29 0.31 2.65 b LOQ
B9 0.63 0.63 2.25 11.28 2.20 0.36 0.65 0.77 1.45 0.48 0.53 0.58 0.48 1.04 0.72
B10 3.88 0.68 1.60 5.29 0.97 0.41 0.66 0.66 0.87 0.56 0.46 0.46 0.46 1.04 0.87
B11 3.88 0.58 1.93 12.53 2.25 0.43 0.68 0.83 1.48 0.50 0.53 0.55 0.53 1.05 0.90
B12 0.44 0.55 0.85 2.45 0.60 0.38 0.65 0.55 0.90 0.50 0.48 0.48 0.45 1.05 0.88
B13 3.49 0.68 1.37 3.56 0.68 0.39 0.66 0.54 0.85 0.49 0.46 0.46 0.71 1.02 0.85
B14 3.70 0.63 3.28 36.85 2.00 12.90 25.65 0.63 0.88 0.50 0.48 0.48 0.45 1.03 0.88
B15 0.48 0.48 2.23 15.35 3.25 0.38 0.68 0.88 1.60 0.48 0.50 0.55 0.55 1.08 0.83
B16 0.69 0.45 2.13 14.69 3.11 0.36 0.65 0.84 1.53 0.45 0.48 0.53 0.29 1.03 0.84
B17 2.06 0.25 1.73 21.58 1.21 1.06 1.74 0.26 2.67 0.74 0.28 0.25 b LOQ bLOQ b LOQ
Mean 2.14 0.40 1.39 9.32 1.34 1.74 8.69 0.51 0.94 0.42 0.34 0.38 0.33 0.94 0.48
(3.60)⁎ (0.24)⁎⁎ (0.97)⁎⁎ (9.54) (1.01)⁎ (3.05) (13.41) (0.30) (0.66) (0.19) (0.18) (0.15)⁎ (0.26) (0.92) (0.40)

b LOQ: less than the limit of quantitation.

Significance: ⁎⁎ (P b 0.01) and ⁎ (P b 0.05).

significant differences (P N 0.05) were observed. An important factor
for PAH contamination is the surface/mass ratio. “Androlla” being of
lesser size and thickness than “Botillo”, showed a larger surface per
unit of volume, which favours adsorption of PAHs.
The PAH values observed in both sausages were clearly lower than
those reported for other smoked meat products (Martí-Cid, Llobet,
Castell, & Domingo, 2008; Purcaro, Moret, & Conte, 2009; Stumpe-
Viksna, Bartkevics, Kukare, & Morozovs 2008) but these values were
higher than those obtained for other Serbian smoked meat products
(Djinovic, Popovic, & Jira, 2008a).
The content of the smoke compounds present in the smoked
products depends on the smoke composition used and on the inten-
sity of deposition on the products of the compounds present in the
smoke. The smoke composition depends on the nature (vegetable
species) of the wood that burns to generate the smoke and on the
temperature to which the wood burns (temperature to which the
thermal decomposition of the wood takes place) (García-Falcón, &
Simal-Gándara, 2005; Muthumbi et al., 2003; Rey-Salgueiro, García-
Falcón, Soto-Gónzalez, & Simal-Gándara, 2004). The intensity of
deposition on the products of the compounds present in the smoke
depends above all on the environmental conditions (temperature and
relative humidity of the air that serves as vehicle for the smoke).
The variability observed by us in the content of each PAH between
the two sausages and also between the units analysed within the
same sausage type comes probably from differences in the type of
wood used and also from differences in the conditions of generation
and deposition of the smoke. The smoking process has been carried
out in a traditional smokestack, in a traditional way, where it is not
possible to control the conditions of the process.
The US Environmental Protection Agency (EPA) has classified
seven PAHs (benzo[a]pyrene, benzo[a]anthracene, chrysene, benzo[b]
fluoranthene, benzo[k]fluoranthene, dibenzo[a,h]anthracene, and
indeno[1,2,3-cd]pyrene) as Group B2, probable human carcinogens
(US Environmental Protection Agency, US EPA, 2002). In turn, the
International Agency for Research on Cancer (IARC) established that
benzo[a]anthracene and benzo[a]pyrene are probable human carci-
nogens, whereas benzo[b]fluoranthene, benzo[k]fluoranthene,
dibenzo[a,h]anthracene, and indeno[1,2,3-cd]pyrene are possible
human carcinogens (IARC, 2004). The total amount of the seven car-
cinogenic PAHs in “Androlla” and “Botillo” samples were 3.93 µg/kg
and 3.40 µg/kg, respectively. If we consider only probable carcino-
genic PAHs, chrysene had the highest content (from 1.782 µg/kg in
“Androlla” samples to 2.67 µg/kg in “Botillo” samples) and was within
the range of those observed by other authors in other smoked meat

Table 2
PAH contents (µg/kg dry weight) in “Androlla” samples.

Code Naph Ace Fln Phe Ant Flt Pyr B[a]A Chr B[b]F B[k]F B[a]P DB[ah]A B[ghi]P I[cd]P

A1 13.53 1.691 3.995 17.770 1.422 0.368 7.892 1.324 1.176 0.735 0.466 0.588 b LOQ bLOQ b LOQ
A2 1.24 0.461 0.874 4.005 0.801 0.388 0.777 0.704 0.898 0.461 0.461 0.461 0.437 0.995 0.850
A3 1.46 0.728 3.786 10.704 2.039 0.607 1.311 0.583 0.898 0.461 0.461 0.485 b LOQ 1.019 0.850
A4 0.92 0.569 5.644 27.475 5.693 1.881 1.535 0.990 1.782 0.470 0.495 0.594 0.495 1.064 0.916
A5 7.43 0.697 6.178 25.409 5.313 0.409 0.625 0.553 0.889 0.481 0.457 0.481 0.433 1.130 b LOQ
A6 0.3 0.493 1.798 4.458 1.010 0.690 1.010 0.542 0.862 0.493 0.468 0.443 b LOQ bLOQ b LOQ
A7 1.36 1.521 3.364 7.258 1.106 0.438 3.387 0.645 0.806 0.599 bLOQ 0.415 0.484 bLOQ b LOQ
A8 7.34 0.597 4.801 22.512 4.328 0.423 0.721 0.721 1.318 0.473 0.473 0.522 0.448 1.169 0.896
A9 2.79 0.558 2.888 9.175 1.820 0.364 0.631 0.534 0.801 0.461 0.461 0.461 0.437 0.995 0.874
A10 7.12 0.488 2.000 6.220 1.244 0.366 0.659 0.537 0.805 0.463 0.463 0.463 0.439 1.024 0.854
A11 10.73 0.510 2.864 10.874 1.966 0.364 0.655 0.534 0.801 0.510 0.461 0.461 b LOQ bLOQ b LOQ
A12 32.9 1.114 12.550 50.050 8.837 0.470 0.792 0.569 0.990 0.495 0.470 0.495 b LOQ 1.040 0.916
A13 10.67 0.597 1.791 3.706 0.697 0.398 0.771 0.547 0.846 0.522 0.473 0.473 0.522 bLOQ b LOQ
A14 4.8 0.481 2.284 10.529 1.875 0.409 0.986 0.673 1.322 0.986 0.505 0.505 0.433 1.010 0.865
Mean 7.32 0.75 3.91 15.01 2.72 0.54 1.55 0.67 1.01 0.54 0.43 0.49 0.29 0.67 0.50
(8.50)⁎ (0.40)⁎⁎ (2.91)⁎⁎ (12.83) (2.40)⁎ (0.40) (1.96) (0.22) (0.28) (0.14) (0.12) (0.05)⁎ (0.23) (0.52) (0.52)

b LOQ: less than the limit of quantitation.

Significance: ⁎⁎ (P b 0.01) and ⁎ (P b 0.05).
 J.M. Lorenzo et al. / Meat Science 86 (2010) 660–664 663

Table 3
Correlational analysis (correlation coefficients, R) of the contents of PAHs found in the investigated “Androlla” samples.

Naph Ace Fln Phe Ant Flt Pyr B[a]A Chr B[b]F B[k]F B[a]P DB[ah]A B[ghi]P I[cd]P ΣPAHs 6 IARC

Naph 1.00
Ace 0.35 1.00
Flt 0.77⁎⁎ 0.35 1.00
Phe 0.73⁎⁎ 0.28 0.97⁎⁎ 1.00
Ant 0.62⁎ 0.09 0.94⁎⁎ 0.97⁎⁎ 1.00
Fln − 0.26 − 0.15 0.18 0.27 0.35 1.00
Py 0.09 0.84⁎⁎ 0.00 0.01 − 0.20 − 0.03 1.00
B[a]A 0.02 0.55⁎ 0.06 0.18 0.03 0.34 0.83⁎⁎ 1.00
Chr − 0.06 − 0.02 0.24 0.41 0.41 0.73⁎⁎ 0.17 0.62⁎ 1.00
B[b]F 0.03 0.26 − 0.14 − 0.09 − 0.20 − 0.18 0.41 0.35 0.29 1.00
B[k]F 0.19 − 0.56⁎ 0.06 0.19 0.21 0.13 − 0.27 0.07 0.29 − 0.04 1.00
B[a]P 0.18 0.20 0.31 0.47 0.39 0.54⁎⁎ 0.46 0.82⁎⁎ 0.84⁎⁎ 0.24 0.47 1.00
DB[ah]A − 0.40 − 0.28 − 0.28 − 0.23 − 0.11 0.13 − 0.29 − 0.10 0.20 0.01 − 0.20 − 0.07 1.00
B[ghi]P 0.01 − 0.40 0.33 0.41 0.51 0.18 − 0.47 − 0.14 0.34 − 0.17 0.38 0.22 0.33 1.00
I[cd]P 0.02 − 0.37 0.21 0.28 0.34 0.24 − 0.38 − 0.05 0.39 − 0.09 0.35 0.24 0.21 0.84⁎⁎ 1.00
ΣPAHs 0.85⁎ 0.37 0.97⁎⁎ 0.98⁎⁎ 0.91⁎⁎ 0.13 0.10 0.20 0.31 − 0.03 0.18 0.44 − 0.30 0.30 0.21 1.00
6 IARC − 0.06 − 0.19 0.07 0.28 0.22 0.34 − 0.01 0.42 0.72⁎⁎ 0.29 0.41 0.63⁎⁎ 0.41 0.68⁎⁎ 0.79⁎⁎ 0.17 1.00

Significance: ⁎⁎ (P b 0.01) and ⁎ (P b 0.05).

products (Djinovic et al., 2008a; Purcaro et al., 2009) but it was lower
than those obtained for salami (Martí-Cid, et al., 2008).
Most of the contents of PAHs found were phenanthrene (9.32 µg/kg),
pyrene (8.69 µg/kg) and naphthalene (2.14 µg/kg) in “Botillo” samples
and phenanthrene (15.01 µg/kg), naphthalene (7.32 µg/kg) and fluorine
(3.91 µg/kg) in “Androlla” samples. In all examined samples content of
phenanthrene was the highest although no significant differences
(P N 0.05) were observed between the two traditional sausage varieties.
Our results agree with those reported by other authors for smoked meat
products (Martí-Cid et al., 2008; Purcaro et al., 2009) since phenan-
threne was the most abundant of PAHs.
The results reported by Jira, Ziegenhals, and Speer (2006) showed
that BaP was the majority compound within the group of other PAHs
with higher carcinogenic potential (Dha, DeP, Dhp; DlP) (Muller et al.,
1995). Reinik, Tamme, Roasto, Juhkam, Tenno, and Kiis (2007) found
that BaP comprised on average 6.1% of the sum of PAHs (BaA, BbF, BkF,
BjF, BgP, BaP, CHR, DhA, Dep, DlP, IcP and 5MC) in commercial smoked
meat products. The contribution of BaP was about 1.2% (mean) and
varied from approximately 0.5 to 5% in “Botillo” meat samples
and about 1.3% (mean) and varied from approximately 1 to 3% in
“Androlla” samples. The BaP values observed in both sausages were
clearly lower than those reported for other smoked meat products
(Djinovic, Popovic, & Jira, 2008b) but these values were similar
with those found in other Spanish sausage (García-Falcón & Simal-
Gándara, 2005) and they were the highest than those obtained for
other Italian smoked sausage (Purcaro et al., 2009).
Recently, the EU has established a maximum level for BaP of 5 μg/kg
wet weight European Commission (2005b) (EC 208/2005) for smoked
meat and smoked meat products although different European
countries (Germany, Austria, Czech Republic and Slovak Republic)
had previously adopted a legal limit of 1 μg/kg. The EU has also set a
maximum permissible limit for BaP present in foodstuffs as a result
of the use of smoking-flavour agents at 0.03 μg/kg (Council of the
European Communities, 1988). In our work any sample did not exceed
5 μg/kg.
Our results showed that BaP was in general 17% and 15% in
“Androlla” meat and “Botillo” meat, respectively of the total sum of
the seven PAHs that are categorized by EPA as probably carcinogenic
to humans.
Toxicological investigations indicated that dibenzo[a,l]pyrene
probably has much stronger carcinogenic potential than benzo[a]
pyrene (Higginbotham, Ramakrishna, Johansson, Rogan, & Cavalieri,
1993) but in this study the content of DlP was not investigated.
The results show that BaP is significantly correlated with Flt, BaA
and Chr, and the sum of PAHs is significantly correlated with Naph,
Fln, Phe and Ant in “Androlla” samples and BaP is significantly
correlated with Db[ah]A and I[cd]P, and the sum of PAHs is sig-
nificantly correlated with Phe, Flt and Pyr in “Botillo” samples
(Tables 3 and 4).
The suitability of BaP as marker for 6 possible and probable
carcinogenic PAHs was also checked by correlation between contents
of BaP and 6 of the total 12 IARC PAH (B[a]A, B[b]F, B[k]F, B[a]P, DB[ah]

Table 4
Correlational analysis (correlation coefficients, R) of the contents of PAHs found in the investigated “Botillo” samples.

Naph Ace Fln Phe Ant Flt Pyr B[a]A Chr B[b]F B[k]F B[a]P DB[ah]A B[ghi]P I[cd]P ΣPAHs 6 IARC

Naph 1.00
Ace 0.25 1.00
Flt 0.56⁎ 0.65⁎ 1.00
Phe 0.30 0.30 0.83⁎⁎ 1.00
Ant 0.31 0.31 0.71⁎⁎ 0.56⁎ 1.00
Fln 0.15 0.04 0.41 0.71⁎⁎ 0.07 1.00
Py − 0.14 − 0.50⁎ − 0.33 − 0.03 − 0.05 0.42 1.00
B[a]A 0.57⁎ 0.70 0.80⁎⁎ 0.39 0.79⁎⁎ − 0.05 − 0.37 1.00
Chr 0.20 0.39 0.63⁎⁎ 0.57⁎ 0.58⁎ − 0.14 − 0.47 0.51⁎ 1.00
B[b]F 0.36 0.40 0.35 0.26 0.31 − 0.12 − 0.06 0.41 0.64⁎⁎ 1.00
B[k]F 0.40 0.91⁎⁎ 0.80⁎⁎ 0.42 0.60⁎ − 0.01 − 0.53⁎ 0.90⁎⁎ 0.59⁎ 0.47 1.00
B[a]P 0.23 0.85⁎⁎ 0.69⁎⁎ 0.32 0.66⁎⁎ − 0.07 − 0.37 0.88⁎⁎ 0.50⁎ 0.54⁎ 0.94⁎⁎ 1.00
DB[ah]A − 0.18 0.52⁎ 0.10 − 0.08 0.03 − 0.11 − 0.30 0.26 0.00 0.41 0.41 0.57⁎ 1.00
B[ghi]P − 0.28 − 0.04 − 0.25 − 0.23 − 0.15 − 0.07 0.11 − 0.09 − 0.24 0.33 − 0.11 0.15 0.71⁎⁎ 1.00
I[cd]P − 0.09 0.86⁎⁎ 0.48 0.23 0.37 0.04 − 0.35 0.59⁎ 0.28 0.38 0.78 0.84⁎⁎ 0.68⁎⁎ 0.19 1.00
ΣPAHs 0.30 − 0.02 0.47 0.72⁎⁎ 0.44 0.82⁎⁎ 0.64⁎⁎ 0.18 0.08 0.20 0.07 0.10 − 0.20 − 0.07 − 0.01 1.00
6 IARC 0.10 0.88⁎⁎ 0.60⁎ 0.28 0.52⁎ − 0.05 − 0.41 0.76⁎⁎ 0.45 0.52⁎ 0.89⁎⁎ 0.96⁎⁎ 0.73⁎⁎ 0.04 0.93⁎⁎ 0.03 1.00

Significance: ⁎⁎ (P b 0.01) and ⁎ (P b 0.05).

664 J.M. Lorenzo et al. / Meat Science 86 (2010) 660–664
A, and I[cd]P) compound contents in both sausages (Tables 3 and 4).
Our results show that BaP is significantly correlated with 6 IARC in
“Androlla” samples (RBaP/6IARC = 0.63; P b 0.01) and in “Botillo” sam-
ples (RBaP/6IARC = 0.96, P b 0.01).
According to ours results, it could be concluded, that BaP is not
good marker for the Σ 15 PAHs that were investigated in this study but
it is a good marker for 6 IARC possible and probable carcinogenic PAHs
in “Androlla” and “Botillo” samples.
4. Conclusions
From the analysis of two Spanish traditional smoked sausages
(“Androlla” and “Botillo”) for contents of PAHs, it can be concluded
that:
• “Androlla” samples showed more contents of PAH compounds than
“Botillo” samples. This circumstance could be attributed to the fact
that “Androlla” has a large surface per unit of volume which favours
adsorption of PAHs.
• Content of phenanthrene was the highest in the two traditional
sausage varieties for all examined samples.
• Differences for PAH content were found between same samples
(obtained by different producer), indicating that conditions used for
smoking can influence the final content of carcinogenic PAHs.
• If we consider only probable carcinogenic PAHs, chrysene showed
the highest content in both traditional sausages.
• The maximum acceptable content of 5 µg/kg for BaP in smoked
meat products was not exceeded in any samples.
• BaP was in general 1.3% and 1.2% in “Androlla” and “Botillo” sam-
ples, respectively of the total sum of the 15 PAHs that were in-
vestigated in this study.
• BaP comprises in general 17% and 15% in “Androlla” and “Botillo”
samples, respectively of the total sum of the seven PAHs that are
categorized by US EPA as probably carcinogenic to humans.
• According to the results from statistical analysis of this study it could
be concluded that BaP could not be used as a marker for 16 EU PAHs
but it is a good marker for 6 IARC possible and probable carcinogenic
PAHs in “Androlla” and “Botillo” samples.
Acknowledgements
This work was a part of the research Project 08TAL009CT granted
by the Xunta de Galicia (The Regional Government). The authors are
grateful to María Figueiredo for technical assistance in HPLC analysis.
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