Silvana Vianna Rodrigues

Polar drug residues in sewage and natural waters in the state of Rio de Janeiro, Brazil.

The drug residues of lipid regulators, anti-inflammatories and some drug metabolites have been detected in raw sewage, treated waste water and river water in the state of Rio de Janeiro, Brazil. These residues are mainly derived from humans via excretion. The median concentrations in the effluents of sewage treatment plants (STPs) of most drugs investigated in this study ranged from 0.1 to 1 microgram/l. The removal rates of individual drugs during passage through a Brazilian STP varied from 12 to 90%. As a consequence of the incomplete removal of these residues during passage through a STP, rivers were also found to be contaminated. Median concentrations ranged from between 0.02 and 0.04 microgram/l in river water, whereas the maximum values were observed to be up to 0.5 microgram/l.

Incomplete degradation of linear alkylbenzene sulfonate surfactants in Brazilian surface waters and pursuit of their polar metabolites in drinking waters.

In Brazil more than 90% of the population are not connected to municipal wastewater treatment plants. As a consequence, surface waters receive continuously considerable amounts of untreated domestic sewage containing surfactants as a major constituent. Such polluted waters gave rise to special interest if they are used as a source for the production of drinking water. In this work, the river Rio Macacu (State Rio de Janeiro, Brazil) was monitored for the occurrence of the most widely used anionic surfactant linear alkylbenzene sulfonate (LAS) together with its main degradative product, sulfophenyl carboxylates (SPC). In order to pursue the fate of both compounds after emission into the river, samples were collected at several locations along the river bank, and analyzed applying liquid chromatography-mass spectrometry after enrichment by solid-phase extraction. The LAS concentrations ranged between 14 and 155 microg l(-1) and the levels of their metabolic intermediates were found from 1.2 to 14 microg l(-1). The self-purification capacity of the water was impressively demonstrated in the upper course of the river downstream of a town considered as one major discharge point, whereas in the lower course the relative constant concentrations of both analytes were detected which was explained with an overall increasing level of pollution. Furthermore, a series of drinking water samples from Niterói and São Gonçalo, supplied by the same waterworks treating surface waters from the Rio Macacu, were taken during two sampling periods and examined for the presence of the strongly polar SPC which is suspected of by-passing the purification processes. The levels detected in the drinking water ranged between 1.6 and 3.3 microg l(-1). For the analyses of drinking and surface waters the peak pattern of a selected SPC homologue composed by several positional isomers served as an indicator to describe the progression of SPC degradation occurred in the river and could be used to distinguish drinking waters of different origins.

TRANSPORT AND DEFENSIVE ROLE OF ELATOL AT THE SURFACE OF THE RED SEAWEED LAURENCIA OBTUSA (CERAMIALES, RHODOPHYTA) 1

Natural within‐thallus concentrations of elatol produced by Laurencia obtusa (Huds.) J. V. Lamour. inhibit herbivory and prevent fouling. However, elatol occurs in larger amounts within the thallus compared with the quantities from the surface of this alga. We evaluated whether the surface elatol concentrations inhibit both herbivory and fouling and whether the content of corps en cerise can be transferred to the external cell walls. Surface elatol concentrations did not inhibit herbivory by sea urchins, settlement of barnacle larvae, or mussel attachment. Evidence of a connection between the corps en cerise, where elatol is probably stored, and the cell wall of L. obtusa was based on channel‐like membranous connections that transport vesicles from the corps to the cell wall region. Therefore, L. obtusa presents a specific process of chemical transport between the cell storage structures and the plant surface. We hypothesized that if high amounts of elatol are capable of inhibiting herbivory and fouling, if the tested organisms are ecologically relevant, and if elatol really occurs on the surface of L. obtusa and this seaweed can transport this compound to its surface, the low natural concentration of defensive chemicals on the surface of L. obtusa is probably not absolute but may be variable according to environmental conditions. We also hypothesized that herbivory and fouling would not exert the same selective force for the production of defensive chemicals on L. obtusa’s surface since the low concentrations of elatol were inefficient to inhibit either processes or distinguish selective pressures.

Inter- and intrapopulation variation in the defensive chemistry of the red seaweed Laurencia dendroidea (Ceramiales, Rhodophyta)

Oliveira A.S., Sudatti D.B., Fujii M.T., Rodrigues S.V. and Pereira R.C. 2013. Inter- and intrapopulation variation in the defensive chemistry of the red seaweed Laurencia dendroidea (Ceramiales, Rhodophyta). Phycologia 52: 130–136. DOI: 10.2216/12-058.1 Variation in the production of defensive chemicals by seaweeds is an important feature to understand the ecological and evolutionary aspects of the predator–prey interactions in marine systems. In this study, we evaluated the inter- and intrapopulation variations in the amount of elatol, which is a sesquiterpene produced as a chemical defence in specimens from four populations of the red seaweed Laurencia dendroidea along the Brazilian coast. The concentration of elatol varied from 2 to 10 times among the L. dendroidea individuals but was approximately three orders of magnitude broader among the populations from 0.001% to 1.24% of the dry weight. The specimens collected at lower latitudes produced low amounts of defences, which did not support the hypothesis that chemical defences increase with a decrease in latitude. When cultivated under distinct temperatures, L. dendroidea specimens produced more defences (= elatol) under high temperatures (25°C). Although we observed a genotypic relevance for chemical defence, the environmental differences along the Brazilian coast support the hypothesis that the variability observed in elatol production by L. dendroidea was most likely influenced by small-scale changes in environmental factors.

Fast and sensitive collagen quantification by alkaline hydrolysis/hydroxyproline assay

A preparative protein alkaline hydrolysis procedure, as part of a spectrophotometric collagen quantification method, is presented. The procedure is suitable for small amounts of fresh solid or liquid samples. Various aspects of the procedure, such as the NaOH concentration, time needed to hydrolyse different collagen contents, buffer strength of the reagent solution, pH control of the hydrolysate and spectrophotometric conditions, were evaluated. Compared to other procedures that use alkaline hydrolysis, the sensitivity of this procedure was increased by a factor of 5. Compared to the conventionally used Association of Official Analytical Chemists (AOAC) acid hydrolysis method, the reaction time was reduced from 16 h to 40 min and the amount of sample from 4 g to 3-20 mg, producing equivalent results when applied to porcine liver and sausage samples.

5 – BIOMASSA FITOPLANCTÔNICA E PRODUÇÃO PRIMÁRIA*

RESUMO A caracterizacao da variabilidade espaco-temporal da biomassa fitoplanctonica e da produtividade primaria da Bacia de Campos foi observada durante duas campanhas oceanograficas realizadas entre marco e abril (periodo chuvoso) e agosto e setembro (periodo seco) de 2009. Durante estas campanhas foram coletadas amostras na superficie e na base da camada de mistura em cinco transectos no periodo chuvoso e quatro no periodo seco, cujas estacoes variaram entre 25 e 3.000 m de profundidade. A concentracao de clorofila a total (mono- e divinil-) apresentou um gradiente decrescente do continente para a regiao oceânica, variando de 0,20 a 1,80 μg·L−1 e 0,10 a 7,83 μg·L−1 na regiao neritica e nos periodos chuvoso e seco, respectivamente. Na regiao oceânica a concentracao variou entre 0,04 e 0,33 μg·L−1 em ambos os periodos do ano. A divinil-clorofila a foi detectada em grande parte da Bacia de Campos com concentracoes variando ate 0,15 μg·L−1 e apresentou padrao de distribuicao inverso ao da clorofila a total, com gradiente longitudinal crescente ao longo da regiao oceânica, chegando a representar 40% da clorofila a total nessas regioes. A produtividade primaria foi avaliada por meio de experimentos in situ simulados para a construcao das curvas de luz-fotossintese e consequente estimativa dos parâmetros fotossinteticos, que foram utilizados para o calculo da produtividade primaria integrada na zona eufotica. Os valores de produtividade primaria variaram de 0,07 a 1,56 gC·m−2·d−1 no periodo chuvoso, e de 0,34 a 0,57 gC·m−2·d−1 no periodo seco, nao havendo diferenca significativa entre os dois periodos. Os resultados obtidos confirmam a caracteristica oligotrofica da regiao da Bacia de Campos, com excecao das regioes proximas de Cabo Frio e Cabo Sao Tome, que estao sujeitas a influencia de feicoes oceanograficas que aportam nutrientes.

Validation of a SPE-LC–MS/MS method for the determination of ketamine and norketamine in micropulverized hair after a single IV dose

A SPE-LC-MS/MS method for the determination of ketamine (KET) and norketamine (NKET) was developed and validated. Extensive pulverization (25min at 25Hz) of previously cooled samples (5min in liquid nitrogen) allowed for extraction in a phosphate buffer (pH 6) solution after 10min vortex agitation at room temperature, simplifying the coupling of the extraction to an effective mixed-mode SPE (solid phase extraction) clean-up procedure. The extraction optimization was performed with samples fortified by drug incorporation according to a published procedure involving incubation of blank matrices for 16days. The method was validated for selectivity, matrix effect, linearity, LLQ (lower limit of quantification), precision, accuracy, recovery, carryover and stability after preparation and has proven to be accurate and reliable within a range of 0.02-10ng/mg for KET and 0.04-4ng/mg for NKET, meeting proposed KET cutoffs for discrimination from chronic use. In addition, the method was sensitive enough to detect the drugs after unique small (1mg/kg) intravenous doses received by patients submitted to general anesthesia before surgical procedures. Ketamine levels varied from 0.060 to 0.111ng/mg and norketamine was positive (<LLQ) in all samples. The high sensitivity achieved with conventional 3D-ion-trap MS/MS detection is probably due to the simple but very effective proposed procedure.

COMPOSIÇÃO DO FITOPLÂNCTON A PARTIR DA AVALIAÇÃO DE PIGMENTOS MARCADORES (CAROTENOIDES E CLOROFILAS)

RESUMO A composicao das comunidades fitoplanctonicas na Bacia de Campos (BC) foi estudada atraves de pigmentos marcadores durante duas campanhas oceanograficas realizadas entre marco e abril (periodo chuvoso – PC) e agosto e setembro (periodo seco – PS) de 2009. Foram coletadas amostras na superficie (1 m) e na base da camada de mistura, em nove transectos (da regiao costeira a oceânica) no PC e no PS. Os pigmentos foram analisados por cromatografia liquida de alta eficiencia (HPLC, sigla em ingles). O software CHEMTAX foi utilizado para estimar, a partir dos pigmentos, a contribuicao dos diferentes grupos fitoplanctonicos para a clorofila a total (TCla). A biomassa fitoplanctonica estimada pela TCla apresentou grande variacao na area da Bacia de Campos, sendo observado um gradiente decrescente da regiao neritica para a regiao oceânica. Os valores observados nas aguas da regiao neritica podem ser considerados tipicos de regioes costeiras (maximo de 1,86 μg·L−1 no PC e 5,51 μg·L−1 no PS). As concentracoes registradas na regiao oceânica sao caracteristicas de sistemas oligotroficos como as aguas da Corrente do Brasil (0,03 a 0,3 μg·L−1 no PC e 0,06 a 0,6 μg·L−1 no PS). A abordagem quimiotaxonomica registrou a presenca de sete classes fitoplanctonicas que apresentaram padroes espaciais ao longo dos perfis batimetrico e vertical, mas nao foram identificados padroes de distribuicao latitudinais. Em geral os grupos fitoplanctonicos tiveram biomassas mais elevadas durante o periodo seco, mas as cianobacterias tiveram as maiores biomassas registradas no periodo chuvoso. Foram observados distintos padroes espaciais no perfil batimetrico. As diatomaceas apresentaram biomassas elevadas nas primeiras isobatas (25 e 50 m), enquanto organismos nanoflagelados (primnesioficeas, prasinoficeas e criptoficeas) mostraram gradiente decrescente ao longo do perfil batimetrico. O picoplâncton eucariotico (pelagoficeas) apresentou gradiente crescente em direcao a quebra da plataforma continental e posterior decrescimo na regiao oceânica. Em relacao as cianobacterias, Trichodesmium (microplâncton), registrado apenas no periodo chuvoso, e Synechococcus (picoplâncton) apresentaram ampla distribuicao em toda Bacia de Campos e maiores biomassas na superficie. Prochlorococcus, outra cianobacteria picoplanctonica registrada, apresentou um padrao semelhante ao das pelagoficeas na campanha do periodo chuvoso, contudo, no periodo seco esteve presente sobretudo na regiao oceânica, apresentando biomassas crescentes a partir da isobata de 150 m. Ao longo do perfil batimetrico pode ser identificado um gradiente da regiao neritica, onde ha maior disponibilidade de nutrientes, para as aguas oceânicas mais oligotroficas, com mudancas na abundância, estrutura de tamanho e composicao taxonomica das assembleias fitoplanctonicas, passando de biomassas mais elevadas do microplâncton para picoplâncton de biomassas reduzidas.

Particle-size distribution (PSD) of pulverized hair: A quantitative approach of milling efficiency and its correlation with drug extraction efficiency

Different types of hair were submitted to different milling procedures and their resulting powders were analyzed by scanning electron microscopy (SEM) and laser diffraction (LD). SEM results were qualitative whereas LD results were quantitative and accurately characterized the hair powders through their particle size distribution (PSD). Different types of hair were submitted to an optimized milling conditions and their PSD was quite similar. A good correlation was obtained between PSD results and ketamine concentration in a hair sample analyzed by LC-MS/MS. Hair samples were frozen in liquid nitrogen for 5min and pulverized at 25Hz for 10min, resulting in 61% of particles <104μm and 39% from 104 to 1000μm. Doing so, a 359% increment on ketamine concentration was obtained for an authentic sample extracted after pulverization comparing with the same sample cut in 1mm fragments. When milling time was extended to 25min, >90% of particles were <60μm and an additional increment of 52.4% in ketamine content was obtained. PSD is a key feature on analysis of pulverized hair as it can affect the method recovery and reproducibility. In addition, PSD is an important issue on sample retesting and quality control procedures.

Using porcine bone marrow to analyze fenproporex and its metabolite amphetamine for forensic toxicological purposes: method development and validation

The possibility of detecting drugs of abuse in bone marrow represents an important issue in forensic toxicology. Bone marrow is an alternative matrix for cases in which the matrices commonly used in forensic practice are unavailable or unfeasible for analysis. The aim of this study was to develop a method for bone cleaning and bone marrow pretreatment and subsequently validate a quantitative method for the simultaneous analysis of fenproporex and amphetamine in porcine bone marrow. Bones were cleaned with proteolytic enzymes and then were cut into small fragments in order to remove the bone marrow. A homogenate was obtained after adding an acidic solution to the bone marrow. Then, the bone marrow homogenate was cleaned with ethyl ether and, after pH adjustment, extracted by liquid–liquid extraction (LLE) with ethyl ether. The dry residue obtained after solvent evaporation was derivatized with N-methyl-bis-trifluoroacetamide (MBTFA)/N-methyl-N-(trimethylsilyl)trifluoroacetamide (MSTFA) and then analyzed by gas chromatography-mass spectrometry (GC-MS). The developed method allowed the identification and quantification of target analytes in the studied matrix. The validation procedure showed good linearity (r2 > 0.99) and recovery (>80%) for fenproporex and amphetamine. The limits of detection and quantitation were 1 ng mg−1 and 5 ng mg−1, respectively, for both analytes. The obtained results are of great importance and may be suitable for potential use in forensic practice.