Nicarbazin Residue in Tissues from Broilers Reared on Reused Litter Conditions.

Nicarbazin (NCZ) is a worldwide used anticoccidial in poultry farming to avoid coccidiosis disease when chickens are reared on conventional poultry litter. If proper dosage and withdrawal time are not followed, the component dinitrocarbanilide (DNC) of NCZ may be present in chicken tissues, therefore posing a risk to consumers if the residues are above 200 µg/kg. Litter reuse is a common and important practice in commercial chicken production. Literature is lacking about the influence of litter reuse on DNC deposition in chicken tissues and its impact on food safety. We aimed to evaluate DNC residues in breast and liver by LC-MS/MS from broilers from an experiment with 10 consecutive flocks during 2 years. The experiment included three treatments containing NCZ in the diet (T1 = 125 mg/kg, 1−21 d; T2 = 125 mg/kg, 1−32 d; T3 = 40 mg/kg, 1−32 d). DNC residues in chicken breast at 21 d in T1 ranged from 648.8−926 µg/kg, at 32 d in T2 and T3 varied, respectively, from 232−667 µg/kg and 52−189 µg/kg. Regarding liver, DNC residues at 21 days in T1 ranged from 11,754−15,281 µg/kg, at 32 days in T2 and T3 varied, respectively, from 10,168−15,021 µg/kg and 2899−4573 µg/kg. When NCZ was withdrawn from feed, DNC residues dropped to

Removal of veterinary antibiotics in swine wastewater using microalgae-based process.

Phycoremediation of swine wastewater is an attractive treatment to remove contaminants and simultaneously produce valuable feedstock biomass. However, there is a lack of information about the application of phycoremediation on veterinary antibiotic removal. Thus, this research investigated the degradation of tetracycline, oxytetracycline, chlortetracycline and doxycycline in swine wastewater treated with phycoremediation. The tetracyclines degradation kinetics was adjusted to the pseudo-first-order kinetics model, with kinetic constant k1 in the following: 0.36 > 0.27>0.19 > 0.18 (d-1) for tetracycline, doxycycline, oxytetracycline and chlortetracycline, respectively. The maximum concentration of microalgae biomass (342.4 ± 20.3 mg L-1) was obtained after 11 days of cultivation, when tetracycline was completely removed. Chlortetracycline concentration decreased, generating iso-chlortetracycline and 4-epi-iso-chlortetracycline. Microalgae biomass harvested after antibiotics removal presented a carbohydrate-rich content of 52.7 ± 8.1, 50.1 ± 3.3, 51.4 ± 5.4 and 57.4 ± 10.4 (%) when cultured with tetracycline, oxytetracycline, chlortetracycline and doxycycline, respectively, while the control culture without antibiotics presented a carbohydrate content of 40 ± 6.5%. These results indicate that could be a valuable source for bioenergy conversion.

Detection of p-Nitroaniline Released from Degradation of 4,4'-Dinitrocarbanilide in Chicken Breast during Thermal Processing.

The diphenylurea 4,4'-dinitrocarbanilide (DNC) is the residue of concern left in edible tissues of broilers fed diets containing the anticoccidial nicarbazin. When chicken meat is submitted to thermal processing, p-nitroaniline (p-NA) is expected from DNC degradation. This work aimed at evaluating whether thermal processing of DNC-containing chicken meat induces p-NA appearance. First, a hydrolysis assay was performed in aqueous solutions at 100 °C in different pH, confirming that DNC cleavage yields p-NA. Then a novel LC-MS/MS method was used to detect traces of this aromatic amine in DNC-containing chicken breast fillets subjected to cooking methods. Our evidence showed p-NA occurrence in such chicken meat samples, which corroborated results from hydrolysis assay. The p-NA appearance in fillets was rather discrete during boiling treatment, but its concentration became pronounced over time for grilling, frying, and roasting, achieving respectively 326.3, 640.0, and 456.9 µg/kg. As far as we are concerned, no other research identified degradation products from DNC residue in heat-processed chicken fillets. Therefore, this study leads to additional approaches to assess impacts on food safety.

Determination of ractopamine residue in tissues and urine from pig fed meat and bone meal.

In many countries, ractopamine hydrochloride (RAC) is allowed to be used in animal production as a ß-agonist, which is an energy repartitioning agent able to offer economic benefits such as increased muscle and decreased fat deposition, feed conversion improvement and an increase in average daily weight gain. However, some countries have banned its use and established strict traceability programmes because of pharmacological implications of ß-agonist residues in meat products. In Brazil, commercial RAC is controlled (5-20 mg kg-1) and only added to pig diet during the last 28 days before slaughter. However, the control is more difficult when co-products, like meat and bone meal (MBM), which can be produced from RAC treated animals, are part of the feed composition. Therefore, a study was undertaken to evaluate the presence of RAC residue concentrations in urine and tissues of gilts (n = 40) in four dietary groups: 0%, 7%, 14% and 21% (w/w) of MBM-containing RAC (53.5 µg kg-1). The concentration of RAC residues in MBM, pig tissues and urine was determined by LC-MS. Low RAC concentrations were detected in muscle, kidney, liver and lungs (limit of detection = 0.15, 0.5, 0.5 and 1.0 µg kg-1, respectively); however, no RAC residues were quantified above the limit of quantification (0.5, 2.5, 2.5 and 2.5 µg kg-1, respectively). In urine, the RAC concentration remained below 1.35 µg L-1. These data suggest that MBM (containing 53.5 µg kg-1 RAC) added to diet up to 21% (w/w) could hamper the trade where RAC is restricted or has zero-tolerance policy.

Application of Enzyme Digestion and Deconjugation Followed by Quick, Easy, Cheap, Effective, Rugged, Safe Extraction and Liquid Chromatography-Tandem Mass Spectrometry Methodology To Determine Ractopamine Residue in Pork.

A new methodology is proposed for ractopamine residue analysis in pork. It consists of enzyme-mediated digestion and deconjugation steps; modified quick, easy, cheap, effective, rugged, and safe (QuEChERS) extraction; and liquid chromatography-tandem mass spectrometry (LC-MS/MS). In brief, the samples were digested with protease and then deconjugated with ß-glucuronidase enzyme; they were then subjected to extraction and cleanup by QuEChERS and underwent sequential analysis by LC-MS/MS. The method performance was evaluated in accordance to the validation guidelines regulated by the Brazilian Ministry of Agriculture and Food Supply. The limit of detection was 0.15 µg/kg and limit of quantification was 0.5 µg/kg. When the method was applied to real samples, ractopamine residue was found in concentrations (up to 7.86 µg/kg) below international recommendation limits up to 10 µg/kg. The method is sensitive, accurate, quick, simple, and suitable for routine analysis; therefore, it is a monitoring tool that may be adopted by laboratories to achieve compliance levels.

Ractopamine analysis in pig kidney, liver and lungs: A validation of the method scope extension using QuEChERS as a sample preparation step.

Ractopamine has been allowed by some countries as a repartitioning additive in pig diet, since it promotes protein synthesis and fat lipolysis. Most regulatory agencies only propose the ractopamine assessment in meat, kidney, liver and fat. Aiming at contributing to the scarcity data regarding this analyte in pig lungs, we extended the scope of a LC-MS method to evaluate pig offals. Homogenized tissue samples were extracted by a QuEChERS procedure; following by clean up steps and further tandem mass spectrometry determination. Method performance was evaluated through specificity, recovery, linearity, reproducibility, repeatability, decision limit (CCα), and detection capability (CCß), in accordance to the Commission Decision 2002/657/EC. Regression coefficients (R2) between 0.994 and 0.999 were achieved for kidney, liver and lungs. Recoveries ranged from 92.0 to 127%. CCα and CCß values ranged from 3.65 to 4.86 µg kg-1, and from 6.27 to 7.21 µg kg-1, respectively. These values were under the maximum residue limits suggested by Codex Alimentarius, which are 90 and 40 µg kg-1 for kidney and liver, respectively. When applied to real samples up to 22.5, 92 and 1003 µg kg-1 of ractopamine residues were detected in pig liver, kidney and lungs, respectively. The results allowed concluding that the proposed analytical method is capable to detect ractopamine residues in all evaluated matrices. Therefore, it can be successfully applied and used as a routine method in laboratories of residue analysis.

Development of a readily applied method to quantify ractopamine residue in meat and bone meal by QuEChERS-LC-MS/MS.

A QuEChERS method of ractopamine (RCT) residue detection in swine meat and bone meal (MBM) samples was demonstrated. Samples were hydrolyzed with protease and ß-glucuronidase prior to QuEChERS (Quick, Easy, Cheap, Effective, Rugged and Safe) extraction and clean-up. Samples were analyzed in a Liquid Chromatography (equipped with ACE 5 C18 column under gradient elution) coupled with a triple quadrupole mass spectrometer operating in positive electrospray ionization mode (using multiple reaction monitoring, MRM). The method was validated for its specificity, decision limit (CCα), detection capability (CCß), recovery, repeatability, reproducibility, linearity, limits of detection (LODs), quantification (LOQs), and stability according to international guidelines (European Commission Decision 2002/657/EC). Recoveries ranged from 96.3 to 107.0%. Repeatability and reproducibility showed both RSD<5.7% and 3.1%, respectively. LODs and LOQs were 1.91 and 6.36 ppb, respectively. CCα and CCß values were 1.91 and 2.37 ppb, respectively. RCT showed good stability for spiked samples and real samples when the concentration was higher, otherwise at lower concentration stability was lower. The proposed method can be successfully applied on a regular basis for the determination of RCT in MBM, demonstrating the usefulness of the method as a tool for compliance monitoring in regulatory laboratories.

Characterization of volatile composition of Laurencia dendroidea by gas chromatography-mass spectrometry

In this study we report the characterization of the volatile compounds of Laurencia dendroidea. Solvent extracts (dichloromethane and methanol), hydrodistillation extracts and headspace solid-phase microextraction samples were obtained and analyzed by GC-MS. Forty-six volatile components were identified in L. dendroidea, among them hydrocarbons, alcohols, phenols, aldehydes, ketones, acids, esters and terpenes.

Biochemical composition of two red seaweed species grown on the Brazilian coast.

BACKGROUND: Algae species have been used as an important source of food because they are highly nutritive considering their vitamin, protein, mineral, fiber, essential fatty acid and carbohydrate contents. However, a large number of seaweeds have been poorly studied, especially Brazilian species. Two red macroalgae species from the Brazilian coast (Plocamium brasiliense and Ochtodes secundiramea) were assessed with respect to their total lipid, fatty acid, total nitrogen, protein, amino acid and total carbohydrate contents. RESULTS: The total lipid contents (dry weight) were 36.3 and 35.4 g kg(-1); fatty acid contents were 9.3 and 12.1 g kg(-1); total nitrogen contents were 37.4 and 24.9 g kg(-1); protein contents were 157.2 and 101.0 g kg(-1); amino acid contents were 127.5 and 91.4 g kg(-1); and total carbohydrate contents were 520.3 and 450.7 g kg(-1) for P. brasiliense and O. secundiramea, respectively. CONCLUSION: Considering these compositions, both algae species were determined to have sources of protein, essential amino acids and carbohydrates similar to the edible seaweeds Laminaria japonica and Palmaria palmata.

Sesquiterpenes from the essential oil of Laurencia dendroidea (Ceramiales, Rhodophyta): isolation, biological activities and distribution among seaweeds

Two known sesquiterpenes (1R*,2S*,3R*,5S*,8S*,9R*)-2,3,5,9-tetramethyltricyclo[6.3.0.0(1,5)]undecan-2-ol and (1S*,2S*,3S*,5S*,8S*,9S*)-2,3,5,9-tetramethyltricyclo-[6.3.0.0(1,5)]undecan-2-ol were isolated for the first time from the essential oil of the red seaweed Laurencia dendroidea collected in the Brazilian coast. These compounds were not active against eight bacteria strains and the yeast Candida albicans, but showed some antioxidant activity. Both compounds were also found in other seaweed species showing that they are not exclusive taxonomic markers to the genus Laurencia.

Composição química e potencial biológico das algas vermelhas marinhas Laurencia filiformis, Laurencia intricata, Plocamium brasiliense e Ochtodes secundiramea da costa brasileira / Chemical composition and biological potency of the marine red algae Laurencia filiformis, Laurencia intricata, Plocamium brasiliense and Ochtodes secundiramea of the Brazilian coast

O oceano apresenta uma vasta diversidade de espécies, entre elas as algas marinhas, as quais são usadas principalmente como fonte de alimentos, de produtos industriais e para uso medicinal. Considerando a biodiversidade encontrada, são poucos os estudos que verificam a composição química e atividade biológica de algas. Desta forma, o presente trabalho descreve especialmente compostos do metabolismo primário (lipídios, proteínas e aminoácidos), composição química volátil, e potencial antioxidante e antimicrobiano de quatro espécies de algas vermelhas da costa brasileira (Laurencia filiformis, Laurencia intricata, Plocamium brasiliense e Ochtodes secundiramea). As análises de lipídeos revelaram que estas algas são ricas em ácidos graxos poliinsaturados ω3 e ω6, mas que apresentam o ácido palmítico como majoritário. O teor de proteínas encontrado é considerável e aproximadamente 50% da composição de aminoácidos é de aminoácidos essenciais. Para extrair os compostos voláteis das algas selecionadas para o estudo, três métodos foram utilizados: arraste a vapor, extração por solvente e HS-SPME. A caracterização química dos compostos voláteis deu-se principalmente pela utilização de cromatografia gasosa acoplada à espectrometria de massas (CG-EM). Ainda foram isolados e identificados dois compostos majoritários do óleo essencial de L. filiformis, o (-)-7-epi-silfiperfolan-6β-ol e o (-)-silfiperfolan-7β-ol, e quatro compostos do extrato acetona/água de P. brasiliense, o 3,4-eritro-7-diclorometil-3-metil-3,4,8-tricloro-1,5(E),7(E)-octatrieno; o 3,4-eritro-7-diclorometil-3-metil-3,4,8-tricloro-1,5(E),7(Z)-octatrieno; o 3,4-eritro-1-bromo-7-diclorometil-3-metil-3,4,8-tricloro-1(E),5(E),7(E)-octatrieno e o 3,4-eritro-1-bromo-7-diclorometil-3-metil-3,4,8-tricloro-1(E),5(E),7(Z)-octatrieno, utilizando diferentes técnicas cromatográficas, como CCDP e CLAE, para isolamento, e técnicas espectroscópicas (RMN uni e bidimensionais) e espectrométricas (HRMS e EIMS) para análise...

The ocean provides large diversity of species, among them the seaweeds, which are mainly used as food, industrial products and as medicine. Considering the biodiversity, there are only few studies which analize the algae volatile compounds and their biological activity. So that, this work describes specially compounds from the primary metabolism (lipids, proteins and amino acids), chemical volatile composition, and antioxidant and antimicrobial potencies of four red algae of the Brazilian coast (Laurencia filiformis, Laurencia intricata, Plicamium brasiliense and Ochtodes secundiramea). The lipid analysis showed that these algae have ω3 and ω6 polyunsaturated fatty acids, but the palmitic acid is the most abundant. The protein content observed is considerable and approximately 50% of the amino acid composition is of essential amino acids. To extract the volatile organic compounds from the algae selected for this study, three methods were used: hydrodestilation, solvent extraction and HS-SPME. For chemical characterization of the volatile compounds, the technique used was gas chromatography coupled with mass spectrometry (GC-MS). In addition, the two most abundant compounds from the essential oil of L. filiformis, the (-)-7-epi-silphiperfolan-6β-ol and the (-)-silphiperfolan-7β-ol, and four compounds of the aceton/water extract of P. brasiliense the 3,4-erythro-7-dichloromethyl-3-methyl-3,4,8-trichloro-1,5(E),7(E)-octatriene; the 3,4-erythro-7-dichloromethyl-3-methyl-3,4,8-trichloro-1,5(E),7(Z)-octatriene; the 3,4-erythro-1-bromo-7-dichloromethyl-3-methyl-3,4,8-trichloro-1(E),5(E),7(E)-octatriene and the 3,4-erythro-1-bromo-7-dichloromethyl-3-methyl-3,4,8-trichloro-1(E),5(E),7(Z)-octatriene, were isolated and identified using different chromatographic techniques like preparative TLC and HPLC for isolation and spectroscopic (NMR uni and bidimensional) and spectrometric techniques (HRMS and EIMS) for analysis. The antioxidant activity of the essential oils, of...

Quinolone alkaloids from Waltheria douradinha.

A phytochemical investigation of the stems of Waltheria douradinha resulted in isolation of two 4-quinolone alkaloids, waltherione B and vanessine, along with three known alkaloids, waltherione A, antidesmone and O-methyltembamide. Their structures were elucidated on the basis of their 2D NMR spectroscopic analyses, and from X-ray crystallographic analysis of waltherione A and the O-methyl derivative of waltherione B. Additionally, waltherione B and vanessine, and the O- and N-methyl derivatives of waltherione A and waltherione B, were evaluated for their antimicrobial activities; only vanessine displayed any (weak) antimicrobial activity.