Environ Sci Pollut Res (2014) 21:3427–3434

DOI 10.1007/s11356-013-2286-5

RESEARCH ARTICLE

Daily ingestion of tetracycline residue present in pasteurized

milk: a public health problem
Sergio Augusto de Albuquerque Fernandes &
Ana Prudencia Assis Magnavita &
Sibelli Passini Barbosa Ferrao &
Simone Andrade Gualberto & Amanda Santos Faleiro &
Abdias Jose Figueiredo & Soraia Vanessa Matarazzo

Received: 21 June 2013 / Accepted: 24 October 2013 / Published online: 17 November 2013
# Springer-Verlag Berlin Heidelberg 2013

Abstract The objective of this study was to evaluate (quali- Keywords Antibiotic residue ingestion . Delvotest® SP-NT .
tatively and quantitatively) the occurrence of antibiotic residue Oxytetracycline . Tetracycline
in pasteurized milk in Brazil. Pasteurized milk samples (n =
252) were collected monthly from Nov. 2010–Oct. 2011 from
21 commercial establishments (brands). A screening test
(Delvotest® SP-NT) was applied to those samples. In positive Introduction
(n =19) and/or suspect samples (n =24), we quantified oxy-
tetracycline (OTC) and tetracycline (TC) by high- In Brazil, monitoring antibiotic residue in milk is a require-
performance liquid chromatography with diode array detector ment according to the official norms for production, process-
(HPLC-DAD). OTCs were detected in all positive samples ing, and commercialization of milk (BRASIL 2003, 2011),
and TCs in six. In the 24suspected samples, OTCs were and meeting the international norms (CODEX
detected in 23 and TCs were not found in 8. Of the milk ALIMENTARIUS 2012) is one of the technological concerns
brands evaluated (n =21), the presence of antibiotic residue of companies in the dairy sector and international public
was not detected in 4; in the other brands, both positive and health.
suspect samples were verified. Results indicate the presence of Milk is a food of great social and economic importance,
antibiotic residue above legal limits. According to actual milk being produced in different edaphoclimatic conditions around
consumption in Brazil (441 mL/kg BW/day), in only 9 of the the world in diverse production systems, from highly to barely
17 brands of milk with antibiotic residue, the estimated daily technical. Its average composition consists of 87.4 % water,
intake was at or less than the maximum recommended by the 12.6 % total solids, 3.9 % fat, 3.2 % crude protein, 4.6 %
European Union. The screening test used was effective to lactose, and 0.9 % minerals. It also includes other solids,
identify the presence of antibiotic residue (OTC and TC), which vary according to species, breed, diet, and other factors.
confirmed by HPLC-DAD. The OTC is the predominant These features indicate its strategic importance in human
antimicrobial used by dairy farmers. Ingestion of contaminat- nutrition, especially for children and the elderly.
ed milk by OTC and TC can increase the resistance of micro- The somatic cell count (SCC) in milk is indicative of the
organisms to antibiotics. health of the mammary gland (MG). Thus, health of the MG
provides milk with less than 105 somatic cells/mL−1 on aver-
Responsible editor: Philippe Garrigues age, considering 2×105 cells/mL−1 as the upper limit for a
health of the MG (Dohoo and Leslie 1991). Higher values of
S. A. de Albuquerque Fernandes (*) : A. P. A. Magnavita :
S. P. B. Ferrao : S. A. Gualberto : A. S. Faleiro : A. J. Figueiredo : the SCC in the MG indicate mastitis. Mastitis negatively
S. V. Matarazzo influences the composition of the milk, which in turn affects
Southwest State University of Bahia, Itapetinga, Bahia, Brazil its processing (Barbano et al. 1991).
e-mail: [email protected]
In Brazil, the SCC varies between 1.5×105 and 106 (Mazal
S. V. Matarazzo et al. 2007; Costa Sobrinho et al. 2012), demonstrating high
State University of Santa Cruz, Ilheus, Bahia, Brazil rates of mastitis infection in cow herds. These results indicate
3428 Environ Sci Pollut Res (2014) 21:3427–3434

the use of medications for treatment of cows and may result in causes losses in the processing of yogurt, butter, and cheese
residue and antibiotics in the milk. due to inhibition of lactic bacteria activity (Knappstein et al.
According to Fonseca et al. (2009) and Oliveira et al. 2003), which makes it necessary to monitor the presence of
(2012), samples positive for antibiotics residue in milk based antibiotic residue in milk (Ilić et al. 2012).
on a screening test range from 0 to 4 % in Brazil. Bando et al. Although Brazil is the fifth largest milk producer in the
(2009) observed that 41.5 % of samples were positive using world (33.2 million tons) (FAO 2013), the rate of dairy prod-
enzymatic tests. Freitas et al. (2013) found 20 % using the LC- ucts for exportation is low. Consequently, improving the milk
IT-TOF MS/QuEChERS; Martins-Júnior et al. (2007) report- quality and offering a safer product is important in increasing
ed positives samples at 8.0 %, using LC-ESI/MS/MS. export. The aim of this study is to qualitatively (Delvotest ®
Antibiotics are used prophylactically or therapeutically in SP-NT) and quantitatively (high-performance liquid chroma-
dairy cattle, especially to control mastitis. This is the principal tography with diode array detector; HPLC-DAD) evaluate the
use of antibiotics in lactating cows as it is the most prevalent presence of antibiotic residue (oxytetracycline, OTC and tet-
disease in these animals (Bansal et al. 2011). The maximum racycline, TC) in pasteurized milk samples to estimate the
residue level (MRL) for tetracyclines classes (oxytetracy- average daily ingestion of this residue by humans.
clines, tetracyclines, and chlortetracyclines) is 100 ng mL−1,
single or in combination (Codex Alimentarius 2012).
The assessment of antimicrobial residue in milk is per- Material and methods
formed using qualitative screening tests in the industry. The
most common test is the inhibition of microbial growth Sampling
(Hennart and Faragher 2012). Several methods are marketed,
including the Delvotest® SP-NT, CMT COPAN®, Eclipse® Analyses were performed in duplicate on 21 brands of
100, Delvotest® Accelerator, and Charm Cowside®. The pasteurized milk. Milk samples (n =252) were collected
Delvotest® SP-NT is reliable, with its broad spectrum of monthly in commercial establishments from November
microbial inhibition in the detection of antibiotic residue; it 2010 to October 2011 in the south and southwest regions
is also easy to use and generates results in a short period of of Bahia State, the largest producer in the States. These
time (3 h), which is essential in the dairy industry (Linage samples were collected in the commercial market (pack-
et al. 2007). The Delvotest® SP-NT (DSM Food Specialties aged in 1,000 mL). After collection, the samples were
Ingredients, Netherlands) allows for detection of five classes frozen and sent to the laboratory for processing of milk
of antimicrobial substances as follows: beta-lactam antibi- and dairy products at the State University of Southwest
otics, sulfonamides, tetracyclines, macrolides, aminoglyco- Bahia (Universidade Estadual do Sudoeste da Bahia),
sides, and others such as trimethoprim and dapsone. where they were identified and remained frozen until
For identification and quantification of antimicrobial resi- analysis.
due present in animal-derived foods, more sensitive and spe-
cific analytical techniques are recommended, such as high- Screening test
performance liquid chromatography (HPLC) (Mamani et al.
2009). HPLC is a fast and reliable technique, with high The Delvotest® SP-NT kit (DSM Food Specialties) was used
sensitivity for the analysis of antibiotic residue like tetracy- as a screening test, and three experts performed this measure-
clines. It permits the confirmation, identification, and quanti- ment visually for the final determination. Samples with posi-
fication of antimicrobial compounds present in the sample tive and/or suspect results were also analyzed by HPLC-DAD.
(Goto et al. 2005). However, some methodologies, such as
liquid chromatography-tandem mass spectrometry (Freitas Sample preparation for extraction
et al. 2013) and ultra-performance liquid chromatography
combined with time-of-flight mass spectrometry (Kennedy Extractions of TCs were performed according to the validated
et al. 1998; Martins-Júnior et al. 2007; Stockler et al. 2009), methodology described by Denobile and Nascimento (2004).
are more efficient to identify and quantify the veterinary drugs Aliquots of 1.0 mL were taken from each milk sample,
in milk rather than liquid chromatography–diode array detec- followed by adding 0.35 mL of a trichloroacetic acid solution
tor (Mamani et al. 2009). in acetonitrile at 80 % (w/v). The mixture was agitated (Ika®,
The presence of antibiotic residue in milk may trigger Werkw Gmbh & Co., Germany) for 5 min with subsequent
allergic phenomena, such as toxic effects (Stolker and centrifugation for 15 min at 15,000 rpm. After centrifugation,
Brinkman 2005) and the development of resistance of micro- the supernatant was collected and filtered with the aid of a
organisms to antibiotics (Gao et al. 2012), making dairy syringe through a Durapore membrane filter (0.45 μm pore),
products a source of risk to the population (Can and Çelik and this was stored in an amber vial under refrigeration until
2012). In the industry, the presence of antibiotics in milk the time of analysis.
Environ Sci Pollut Res (2014) 21:3427–3434 3429

Equipment cation by CLAE of these substances in pasteurized milk

samples were (1) linearity data showed a positive correlation
For identification and quantification of tetracyclines in the between the concentrations of tetracyclines (OTC, y=25.2x+
samples, a model SPD20A liquid chromatography was used 49.16 and R2 =0.999; TC, y=31.8x+518.8 and R2 =0.999) in
(Shimadzu, Kyoto, Japan) with a quaternary solvent injection the ranges of 100, 200, 400, 800, and 1,600 ng mL−1. The
valve and a sample loop of 20 μL, column oven, and diode linear correlation coefficients (0.999) found for the curves are
array detector (DAD). The antimicrobial compounds TC and in compliance with those set by ANVISA (2003); (2) the
OTC were separated in a C 8 analytical column (15 cm×4 mm coefficients of variation (CV) obtained in the study of intra/
di × 5 μm). A solution of 0.01 M oxalic acid, acetonitrile, and interday precision, of the milk samples added with the stan-
0.1 % triethylamine (70:20:10 v/v/v) was used as the mobile dards of OTC and TC, in concentrations of 200, 400 and
phase at a flow rate of 1 mL min−1, and the temperature of the 800 ng mL−1, respectively, were 1.7, 2.5, and 1.9 (intraday)
oven was adjusted to 40 °C. The analysis time was 10 min. and 1.7, 3.6, and 4.5 % (interday) for oxytetracycline and 1.7,
The solvents used were chromatographic grade (JC Baker) 2.6, and 1.7 (intraday) and 6.8, 7.5, and 8.1 % (interday) for
and were filtered and degassed before use. The chromato- tetracycline; (3) the detection limits and quantification were of
grams were processed at 363 nm. Identification of OTC and 50 ng mL−1 for OTC and TC, with CV 6.8 and 8.7 %,
TC was tentatively performed by comparing the retention time respectively, and (4) the recovery rate values of tetracyclines
of the peaks with the standard samples, oxytetracycline hy- in the studied concentrations (100, 200, and 400 ng mL−1)
drochloride (95 %) and tetracycline (88 %), both acquired were 64, 64, and 67 % (OTC) and 46, 54, and 61 % (TC) for
from Sigma-Aldrich® Chemical Co. (St. Louis, MO, USA). each concentration, respectively
Evaluation of the presence of antibiotic residue in the milk
samples analyzed was performed by observing the results of Calculation of the estimated daily intake
screening tests (descriptive analysis) and determining the per-
centages of positive, suspect, and negative samples expressed The concentration was determined using the average
in nanograms per milliliter. content of total tetracyclines observed, and the estimated
The general characteristics of HPLC-DAD of the OTC and daily intake (EDI) was calculated according to Juan
TC proved to be effective for the identification and quantifi- et al. (2010).

EDI ¼ ½ðmean in μg of the antibiotic residue per Kg of foodÞ  ðMean consumption of the foodފ
Live adult weight of 60 Kg

For determination of the EDI, the estimated milk consump- suspected samples was greater. Of the 20 positive milk sam-
tion values used were for Brazil in 2012 (441 mL/inhabitant/ ples, 13 (65 %) were detected between January and June. The
day), and this was compared to the recommended consump- total number of suspected samples observed during the year
tion by the Brazilian Ministry of Health, which is 575 mL/ was 25, where 22 (91.7 %) were obtained between January
inhabitant/day (Embrapa 2013). This evaluated the impact of and June (Fig. 1). These results indicate that season likely had
increased milk consumption on the estimate. an effect on the occurrence of positive and suspected milk
samples, as was observed by Kaya and Filazi (2010).
Of the 21 pasteurized milk brands, only four (D, E, P, and
Results T) showed no positive results over the year of study. All other
brands (17) tested positive or suspect at some point (Fig. 2).
Among the 252 pasteurized milk samples analyzed by the Brand N showed the highest number of samples with
Delvotest® SP-NT in this study, 207 (82.0 %) were negative positive (25 %) and suspect (25 %) results, whereas L and O
(no antibiotic residue), and 20 presented positive results, both presented 17 % positive samples and 8 % suspect sam-
revealing a contamination frequency of 8.0 %. On the other ples, respectively. For brands B, C, and M, 17, 17, and 25 % of
hand, 25 samples (10 %) were defined as suspect. In most of the samples were reported as positive, respectively. Brands A,
the milk samples studied, the presence of antibiotic residue F, H, Q, S, and V presented positive samples (8.3 % each) with
(AM) was not detected by the screening test. variations in the percentage of suspect samples. However,
Distribution of the results throughout the year indicated brands G, I, J, R, and U exhibited only suspect samples (25,
that there was a period in which the number of positive and 17, 8, 16.7, and 17 %, respectively).
3430 Environ Sci Pollut Res (2014) 21:3427–3434

Fig. 1 Monthly distribution of

Suspecteds
negative, positive, and suspected 21 21 21
pasteurized milk samples by 19 19 19
Positives

Dairy industry numbers (n = 21)

screening test in dairy industry 18 Negatives
17
(Nov. 2010 to Oct. 2011)
15
14
13

10 10

5
4 44 4

2 2 2 2 2 2
1 1

Jan Feb Mar Apr May Jun Jul Aug Sept Oct Nov Dec
Months

The positive and suspect samples discovered during the (N-Mar). In 16 samples, the presence of TC was not detected;
screening test were subjected to HPLC-DAD for confirmation only 1 sample (N-mar) showed no OTC (Fig. 4).
of the presence of tetracyclines. The antibiotic concentrations observed in most samples
In samples with positive results (n =19), the highest con- with positive or suspect results exceed the MRL permitted
centration observed individually for OTC was 2,782 ng mL−1 for tetracyclines (100 ng mL−1, singly or in combination).
(Fig. 3) in sample I-Jan. In the same sample, the concentration Of the 19 positive samples in the screening tests, all pre-
of TC was 1,145 ng mL−1. The lowest concentrations above sented OTC concentrations above the MRL (Table 1); in
the maximum residue limit (MRL=100 ng mL−1) were ob- six, the TC concentration was excessive; and in one, it was
served in samples F-Apr (125 ng mL−1) for OTC and sample below the legal limit. In 12 samples, TC was not detected.
S-Mar (107 ng mL−1) for TC. In 12 samples, no TC residue Among the samples considered suspect in the screening
was detected, and in one (P-Sept), the TC concentration was test, in 21, the OTC concentration was greater than that
below the MRL (73 ng mL−1), in which no false positive was permitted by legislation; in 2, the concentration was lower;
detected. and in 1, no OTC was detected. TC was not detected in 16
In suspect samples, the lowest OTC concentration was samples; only in 1 did it not exceed the MRL; and in 7, it
observed in sample U-Mar (67 ng mL−1), and the highest exceeded the MRL. The “total tetracycline concentration”
was 2,072 ng mL−1 (S-Feb). With respect to TC, the highest in suspect samples was below the MRL in only three
was 2,297 ng mL−1 (R-Feb), and the lowest was 78 ng mL−1 samples.

Fig. 2 Percentage of negative,

positive, and suspect pasteurized
milk samples by screening test per
dairy industry (Nov. 2010 to Oct.
2011)
Sample percentage

Dairy industry
Environ Sci Pollut Res (2014) 21:3427–3434 3431

Fig. 3 Antibiotics residues 3000

concentration (OTC 2782
2800 OTC
oxytetracycline, TC tetracycline)
2600 TC
by HPLC-DAD in the positives
milk samples in the screening test 2400 2354

(Delvotest® SP-NT). MRL maxi-

Concentração em ng.mL-1
2200
mum residue limit=100 ng mL−1 1926
2000

1800

1600

1400
1145
1200 1051
938 929
1000 848 875 855 885
828 775
800 736 701
685
597
600 506
316 359
400 282
125 135 194
200 73 107 0

0
A-nov B-sept B-nov C-aug C-nov E-sept P-sept F-apr H-jun H-feb I-jan I-feb I-jun Q-jun Q-jul S-may N-feb N-apr N-may
Dairy industry-month

The EDI was based on the average concentration of tetra- the actual milk consumption in Brazil (Fig. 5), it was observed
cycline residue in milk, per dairy facility, observed in this study. that of the 16 brands, in 9, the EDI was in accordance with that
Average daily milk consumption in Brazil in 2011 was recommended by the European Union (EMEA 1995), with
441 mL/inhabitant/day, but the desired average milk con- variations between 1.3 and 3 μg/kg BW/day. In the other 7
sumption is 575 mL/inhabitant/day (Embrapa 2013). When brands, these estimates varied between 3.2 and 8.8 μg/kg BW/
evaluating the EDI of antibiotics (tetracycline) by means of day, where the greatest value was observed in brand I. The

Fig. 4 Concentration (ng/mL−1)

of antibiotics residues (OTC
oxytetracycline, TC tetracycline)
by HPLC-DAD in the suspects
milk samples by screening test
(Delvotest® SP-NT). MRL maxi-
mum residue limit=100 ng mL−1
3432 Environ Sci Pollut Res (2014) 21:3427–3434

Table 1 Number of positive and suspect pasteurized milk samples sold milk). Bando et al. (2009), using immunoassay, found that
according to the screening test for antibiotic residues, confirmed by
41.5 % of samples were positive. Freitas et al. (2013) found
HPLC-DAD
20 % (LC-IT-TOF MS/QuEChERS), and Martins-Júnior et al.
MRL/milk Positive (n =19) Suspect (n =24) (2007) reported 8 % (LC-ESI/MS/MS). In our study, all
positive samples and most suspect samples (87.5 %) exceeded
OTC TC TTC OTC TC TTC
the MRL (Table 1).
Greater than 19 06 19 21 07 21 Screening tests are quick and easy to use, but they are
Less than – 01 – 02 01 03 subjective, so we need more precise and technical measures,
Not detected – 12 – 01 16 – such as liquid chromatography, to find the dynamic of the
contamination of antibiotics in dairy cattle in Brazil. It is
OTC oxytetracycline, TC tetracycline, TTC total tetracyclines necessary improve the sampling number and submit it to
liquid chromatography.
simulation that utilized values of the recommended milk con- The Delvotest SP-NT has low sensitivity to OTC (Le
sumption of 575 mL/inhabitant/day indicated that only 3 of Breton et al. 2007), but in this study, it was effective even at
the 16 brands met the maximum recommended by the low concentrations because it detected even a minimal pres-
European Union for EDI of antibiotics (3 μg/kg BW/day), ence of antibiotic residue in milk (67 ng mL−1) (Fig. 4),
with a variation between 1.7 and 2.9 μg/kg BW/day. In 13, the showing its sensitivity to the analyte (Hennart and Faragher
EDI exceeded the recommendations (3.3 to 11.5 μg/kg BW/ 2012; Sierra et al. 2009).
day). Rainfall in the studied area is concentrated during the
summer season, from December to March (Moscati and Gan
2007). The high humidity could facilitate the development
Discussion of microorganisms that cause mastitis, increasing its inci-
dence. This higher mastitis incidence may increase the de-
Sampling from large volumes can lack analytical precision tection of antibiotic residue (Fig. 1) because mastitis is the
(dilution effect) (FAO 2000), especially during screening tests. most common cause of antibiotic use in lactating cows
In our study, the samples were collected from large volume (Bansal et al. 2011). Milk production systems in the
industries, but the percentage of positive samples was still region involve minimal technology and milking hygiene
close to 20 %. measures are of little use, making these environments fertile
Distinct percentages of pasteurized milk samples positive for mastitis.
for antibiotics after a screening test were reported in Brazil. Our results indicate that within the class of tetracyclines,
Oliveira et al. (2012) observed no antibiotic residue in milk OTC is most commonly used among the dairy products of the
samples (raw and pasteurized) evaluated. Fonseca et al. (2009) region studied as well as in other countries (Schlegelova et al.
found 0–4 % noncompliance of samples evaluated (UHT 2002).

Fig. 5 Estimated daily intake 12.0 BR actual (441 mL/inhabitant/day) 11.5

EDI simulation (μg/kg body BR/MS (575 mL/inhabitant/day)
weight/day) of the antimicrobial 10.4
residue (tetracyclines) in milk.
Recommendation of the EMEA
(1995) of average daily con- 8.8
EDI (µg/Kg Body Weight/day)

9.0
sumption of tetracyclines=3 μg/ 7.9
kg BW/day 7.4 7.4

6.2 6.2
6.0 5.7 5.7

4.7 4.7
4.1
3.9 3.7
3.5 3.5
3.3 3.3 3.2
3.0 2.9 2.9
3.0 2.7 2.5 2.7
2.5
2.2 2.2
1.7 1.7
1.3

0.0
A B C E F G H I L M N P Q R S U
Dairy industry
Environ Sci Pollut Res (2014) 21:3427–3434 3433

The results obtained in this study indicate the need for References
greater monitoring and control by health surveillance agencies
of the production and consumption of milk. One of the major Bando E, Oliveira RC, Ferreira GM, Machinski M Jr (2009) Occurrence
causes of antibiotic residue in milk is animal treatment with of antimicrobial residues in pasteurized milk commercialized in the
antibiotics; such animals should be milked separately for a state of Paraná, Brazil. J Food Prot 72(4):911–914
Bansal BK, Bajwa NS, Randhawa SS, Ranjan R, Dhaliwal PS (2011)
specified period. These substances may be excreted with the
Elimination of erythromicynin in milk after intramammary admin-
milk even after finalizing treatment, making it necessary that istration in cows with specific mastitis: relation to dose, milking
such milk be discarded during this period, which may vary frequency, and udder health. Trop Anim Health Prod 43:323–329
according to the recommendations of the manufacturer Barbano DM, Rasmussen RR, Lynch JM (1991) Influence of milk
somatic cell count and milk age on cheese yield. J Dairy Sci 74:
(Knappstein et al. 2003; Stockler et al. 2009; Bansal et al.
369–388
2011). In addition, it is important for quality control at the Brasil. 2003. National Agency of Sanitary Vigilance. Health Ministry.
dairies to reduce the risks to public health (e.g., toxic effects Resolução RDC N. 119. PAMVET. Diário Oficial da União, 2003;
and development of resistance to pathogenic microorganisms 18 de setembro.
Brasil. (2011) Agricultural, Livestock and Supply Ministry. Normative
in humans) (Gao et al. 2012; Can and Çelik 2012) or to
Instruction, nº 62 de 29 de Dec.-2011. Diário Oficial da União.
animals. Soil contamination is also a problem (Carvalho Brasília, 29 de dezembro de 2011.
et al. 2013). In the dairy industry, antibiotics inhibit lactic Can HY, Çelik TH (2012) Detection of enterotoxigenic and antimicrobial
culture and affect the quality and identity of cheeses by resistant S. aureus in Turkish cheeses. Food Control 24:100–103
Carvalho PN, Pirra A, Basto MCP, Almeida CMR (2013) Activated
altering their composition and structure (Knappstein et al.
sludge systems removal efficiency of veterinary pharmaceuticals
2003). Adjustments to the Brazilian milk production chain from slaughterhouse wastewater. Environ Sci Pollut Res. doi:10.
are imperative; a new development model is fundamental and 1007/s11356-013-1867-7
should be supported by better quality and safety of products Codex Alimentarius. (2012). Codex Alimentarius Commission
Maximum Residue Limits for Veterinary Drugs in Food. Updated
(Farina et al. 2005; Ilić et al. 2012).
as at the 34th Session of the Codex Alimentarius Commission (July
The consumer contamination risk is in a direct relationship 2011).http://www.codexalimentarius.org/committees-task-forces/
with the danger of the chemical product and exposure to the en/?provide=committeeDetail&idList=6. Accessed 04 Oct. 2013.
same (risk = hazard × exposure) (Davies et al. 2003). Costa Sobrinho PS, Faria CAM, Pinheiro JS, Almeida HG, Pires CV,
Antibiotic residue in milk products presents a danger to the Santos AS (2012)Bacteriological Quality of Raw Milk Used for
Production of a Brazilian Farmstead Raw Milk Cheese. Foodborne
consumer, so these results indicate that the inappropriate use Pathog Dis 9(2). doi:10.1089/fpd.2011.1010.
of antibiotics in dairy farming (high dosages, not observing Davies L, O’connor M, Logan S (2003) Chronic intake. In Pesticide
the grace periods) may pose health risks to the public as well Residues in Food and Drinking Water: Human Exposure and
as lead to economic losses in the dairy industry. Risks. John Wiley & Sons Ltd, Chichester
ANVISA - Agência Nacional de Vigilância Sanitária (2003). Resolução
Re. Nº 899, de 29 de Maio de 2003.
Denobile M, Nascimento SE (2004) Method validation for the determi-
Conclusions nation of the antibiotics residues oxytetracycline, tetracycline, chlor-
tetracycline, and doxycycline in milk by high-performance liquid
chromatography. Rev Bras Ciên Farm 40:209–218, in portuguese
The results obtained indicate the presence of antibiotic residue
Dohoo IR, Leslie KE (1991) Evaluation of changes in somatic cell counts
above the legal limits within the class of tetracyclines. OTCs as indicators of new intramammary infections. Prev Vet Med 10:
are the most commonly found in dairy products in the region 225–237
studied. Embrapa—Empresa Brasileira de Pesquisa Agropecuária.
Further studies are required as follows: (1) antimicrobials NationalMilkDiagnostic. 2013. http://www.cnpgl.embrapa.br/nova/
Plano_Pecuario_2012.pdf (in portuguese). Accessed 07 may 2013.
resistance, (2) epidemiological status, (3) effects on the EMEA. (1995). Oxytetracycline, Tetracycline, Chlortetracycline, sum-
ground water, especially near the dairy industry, and (4) mary report (3) committee for veterinary medicinal products.
screening tests followed by HPLC with more sensitive detec- EMEA. EMEA/MRL/023/95.
tors, such as mass spectrometry. It is recommended that train- FAO. (2013). Food and Agriculture Organization for United Nations.
Statistics. Acces 09/12/2013.http://www.fao.org/economic/ess/en/#.
ing and guidance, particularly on the use of proper practices of Ug5-KJLVAZ4.
hygiene during milking, be offered to the owners and their Farina EMMQ, Gutman GE, Lavarello PJ, Nunes R, Reardon T (2005)
workers involved in the dairy industry. Private and public milk standards in Argentina and Brazil. Food
Policy 30:302–315
Fonseca GP, Cruz AG, Faria JAF, Silva R, Moura MRL, Carvalho MRJ
Acknowledgments This research was funded by Fundacao de Amparo
(2009) Antibiotic residues in Brazilian UHT milk: a screening study.
a Pesquisa do Estado da Bahia (FAPESB) and Coordenacao de
Ciênc Tecnol Aliment Campinas 29(2):451–453, abr.-jun. 2009
Aperfeicoamento de Pessoal de Nivel Superior (Capes) for the support
Food and Agriculture Organization of the United Nations & World Health
to the research.
Organization (2000). Codex Alimentarius Commission. CX/RVDF
00/12. Joint Fao/Who Food Standards Programme Codex
Conflict of interest None of the author has any support or relationship Committee on Residues of Veterinary Drugs in Foods. Twelfth
with other people that could inappropriately influence. Session, Washington, D.C.
3434 Environ Sci Pollut Res (2014) 21:3427–3434

Freitas SKB, Paim APS, Silva PTS (2013) Development of a LC-IT-TOF for antimicrobial detection in ovine milk. Jour Dairy Sci 90:5374–
MS procedure to quantify veterinary drug residues in milk 5379
employing a QuEChERS approach. Food Anal Methods. doi:10. Mamani MCV, Reyes FGR, Rath S (2009) Multiresidue determination of
1007/s12161-013-9595-7 tetracyclines, sulfonamides, and chloramphenicol in bovine milk
Gao J, Yu FQ, Luo LP, He JZ, Hou RH, Zhang HQ, Li SM, Su JL, Han B using HPLC-DAD. Food Chem 117:545–552
(2012) Antibiotic resistance of Streptococcus agalactiae from cows Martins-Júnior HA, Kussumi TA, Wang AY, Lebre DT (2007) A rapid
with mastitis. Vet J 194:423–424 method to determine antibiotic residues in milk using Liquid
Goto T, Ito Y, Yamada S, Matsumoto H, Oka H (2005) High-throughput Chromatography Coupled to Electrospray Tandem Mass
analysis of tetracycline and penicillin antibiotics in animal tissues Spectrometry. J Braz Chem Soc 18(2)397–405
using electrospray tandem mass spectrometry with selected reaction Mazal G, Vianna PCB, Santos MV, Gigante ML (2007) Effect of somatic
monitoring transition. J Chromatogr A 1100:193–199 cell count on Prato cheese composition. J Dairy Sci 90:630–636
Hennart, S. T. A.; Faragher, J. 2012. Validation of Delvotest SP NT Moscati MCL, Gan MA (2007) Rainfall variability in the rainy season of
DA.JAOAC, 95, 252. semiarid zone of Northeast Brazil (NEB) and its relation to wind
Ilić K, Jakovljević E, Škodrić-Trifunović V (2012) Social–economic regime. Int J Climatol 27:493–512
factors and irrational antibiotic use as reasons for antibiotic resis- Oliveira LP, Barros LSS, Silva VC, Cirqueira MG (2012)
tance of bacteria causing common childhood infections in primary Microbiological quality and detection of antibiotic residue in raw
healthcare. Eur J Pediatr 171:767–777 and pasteurized milk consumed in the Reconcavo Area of the State
Juan C, Moltó JC, Mañes J, Font G (2010) Determination of macrolide of Bahia, Brazil. Food Process Technol 3(1):2–5. doi:10.4172/2157-
and lincosamide antibiotics by pressurized liquid extraction and 7110.1000137
liquid chromatography tandem mass spectrometry in meat and milk. Schlegelova J, Babák V, Klímová E, Lukásová J, Navrátilová P,
Food Control 21:1703–1709 Sústácková A, Sédivá I, Rysánek D (2002) Prevalence of and
Kaya SE, Filazi A (2010) Determination of antibiotic residues in milk resistance to anti-microbial drugs in selected microbial species iso-
samples. Kafkas Univ Vet Fak Derg 16(Suppl-A):S31–S35 lated from bulk milk samples. J Vet Med B 49:216–225
Kennedy DG, McCracken RJ, Cannavan A, Hewitt SA (1998) Use of Sierra D, Contreras A, Sánches A (2009) Short-communication:
liquid chromatography–mass spectrometry in the analysis of resi- detection limits of non-beta-lactam antibiotics in goat's milk
dues o antibiotics in meat and milk. J Chromatogr A 812:77–98 by microbiological residues screening tests. J Dairy Sci 92:
Knappstein K, Suhren G, Walte GH (2003) Influence of milking frequen- 4.200–4.206
cy on withdrawal period after application of beta-lactam antibiotic- Stockler RM, Morin DE, Lantz RK, Constable PD (2009) Effect of
based drugs. Anal Chim Acta 48(3):241–249 milking frequency and dosing interval on the pharmacokinects of
Le Breton MH, Savoy-Perroud MC, Diserens JM (2007) Validation and cephapirin after intramammary infusion in lactating cows. J Dairy
comparison of the Copan milk test and Delvotest SP-NT for the Sci 92:4262–4275
detection of antimicrobials in milk. Anal Chim Acta 586:280–283 Stolker AAM, Brinkman UA (2005) Analytical strategies for residue
Linage B, Gonzalo C, Carriedo JA, Asensio JA, Blanco MA, La Fuente analysis of veterinary drugs and growth-promoting agents in food-
LF, Primitivo FS (2007) Performance of blue-yellow screening test producing animals—a review. J Chromatogr Anal 1067:15–53