Instituto Nacional Of Saude Doutor Ricardo Jorge Insa

Porto, Portugal

Instituto Nacional Of Saude Doutor Ricardo Jorge Insa

Porto, Portugal
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Andre C.,University of Lisbon | Delgado I.,Instituto Nacional Of Saude Doutor Ricardo Jorge Insa | Castanheira I.,Instituto Nacional Of Saude Doutor Ricardo Jorge Insa | Bordado J.,University of Lisbon | Matos A.S.,New University of Lisbon
Acta IMEKO | Year: 2017

A statistical experimental design was used to screen variables of the analytical procedure to quantify free monomeric isocyanates presented in polyurethane based pre-polymers in trace amounts. For this purpose, diphenylmethane-4,4'-diisocyanate (4,4'-MDI), 2,4-Toluene diisocyanate (2,4-TDI) and 2,6-Toluene diisocyanate (2,6-TDI) were analysed by Ultra Performance Liquid Chromatography with a Photo Diode Array detector (UPLC-PDA). A preliminary study was performed with three derivatization agents, being 1-(2-piridyl) piperazine (1,2-PP) the most suitable one. Column temperature, flow and percentage of ammonium acetate (% NH4Ac.) were the factors studied at two levels each. A sequence of experiments was planned according to a 23 full factorial design with three replicates and two repetitions. Analysis of variance (ANOVA) was applied for the identification of significant factors and interactions. Higher responses were achieved when the column temperature was 30 °C, a flow of 0.3 mL min-1 and a solvent with a percentage of ammonium acetate of 0.1 %. Figures of merit were assessed within-laboratory as a preliminary step for method validation. Similar values were obtained for TDI and MDI. Recoveries are approximately 100 %. In addition, the values of detection limits (LODs) for MDI and TDI were 0.08 and 0.11 μg mL-1, respectively, and quantification limits (LOQs) were 0.25 and 0.33 μg mL-1 for MDI and TDI, respectively. The working range was between 0.01 and 10.00 μg mL-1 for MDI and 0.01 - 4.95 μg mL-1 for TDI. These figures of merit seemed adequate to detect low amounts of free monomeric isocyanates presented in agglomerates and foams for agglomerated cork stoppers production. This data is suitable to address the optimization of an analytical method by a response surface methodology. © 2017 IMEKO.


Pereira T.,Abel Salazar Biomedical Sciences Institute | Pereira T.,Institute Ciencias e Tecnologias Agrarias e Agro Alimentares ICETA | Gartner A.,Institute Ciencias e Tecnologias Agrarias e Agro Alimentares ICETA | Amorim I.,Abel Salazar Biomedical Sciences Institute | And 21 more authors.
BioMed Research International | Year: 2014

In peripheral nerves MSCs can modulate Wallerian degeneration and the overall regenerative response by acting through paracrine mechanisms directly on regenerating axons or upon the nerve-supporting Schwann cells. In the present study, the effect of human MSCs from Wharton's jelly (HMSCs), differentiated into neuroglial-like cells associated to poly (DL-lactide-ε-caprolactone) membrane, on nerve regeneration, was evaluated in the neurotmesis injury rat sciatic nerve model. Results in vitro showed successful differentiation of HMSCs into neuroglial-like cells, characterized by expression of specific neuroglial markers confirmed by immunocytochemistry and by RT-PCR and qPCR targeting specific genes expressed. In vivo testing evaluated during the healing period of 20 weeks, showed no evident positive effect of HMSCs or neuroglial-like cell enrichment at the sciatic nerve repair site on most of the functional and nerve morphometric predictors of nerve regeneration although the nociception function was almost normal. EPT on the other hand, recovered significantly better after HMSCs enriched membrane employment, to values of residual functional impairment compared to other treated groups. When the neurotmesis injury can be surgically reconstructed with an end-to-end suture or by grafting, the addition of a PLC membrane associated with HMSCs seems to bring significant advantage, especially concerning the motor function recovery. © 2014 T. Pereira et al.


David S.,Instituto Nacional Of Saude Doutor Ricardo Jorge Insa | Mateus A.R.A.,Instituto Gulbenkian Of Ciencia Igc | Mateus A.R.A.,Leiden University | Duarte E.L.,University of Évora | And 5 more authors.
PLoS ONE | Year: 2015

Major contributions from pathogen genome analysis and host genetics have equated the possibility of Mycobacterium tuberculosis co-evolution with its human host leading to more stable sympatrichost-pathogen relationships. However, the attribution to either sympatric or allopatric categories depends on the resolution or grain of genotypic characterization. We explored the influence on the sympatric host-pathogen relationship of clinical (HIV infection and multidrug-resistant tuberculosis [MDRTB]) and demographic (gender and age) factors in regards to the genotypic grain by using spacer oligonucleotide typing (spoligotyping) for classification of M. tuberculosis strains within the Euro-American lineage. We analyzed a total of 547 tuberculosis (TB) cases, from six year consecutive sampling in a setting with high TB-HIV coinfection (32.0%). Of these, 62.0% were caused by major circulating pathogen genotypes. The sympatric relationship was defined according to spoligotype in comparison to the international spoligotype database SpolDB4. While no significant association with Euro-American lineage was observed with any of the factors analyzed, increasing the resolution with spoligotyping evidenced a significant association of MDRTB with sympatric strains, regardless of the HIV status. Furthermore, distribution curves of the prevalence of sympatric and allopatric TB in relation to patients' age showed an accentuation of the relevance of the age of onset in the allopatric relationship, as reflected in the trimodal distribution. On the contrary, sympatric TB was characterized by the tendency towards a typical (standard) distribution curve. Our results suggest that within the Euro-American lineage a greater degree of genotyping fine-tuning is necessary in modeling the biological processes behind the host-pathogen interplay. Furthermore, prevalence distribution of sympatric TB to age was suggestive of host genetic determinisms driven by more common variants. © 2015 David et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.


Dias M.G.,Instituto Nacional Of Saude Doutor Ricardo Jorge Insa | Oliveira L.,Instituto Nacional Of Saude Doutor Ricardo Jorge Insa | Camoes M.F.G.F.C.,University of Lisbon | Nunes B.,Instituto Nacional Of Saude Doutor Ricardo Jorge Insa | And 2 more authors.
Journal of Chromatography A | Year: 2010

Three sets of extraction/saponification/HPLC conditions for food carotenoid quantification were technically and economically compared. Samples were analysed for carotenoids α-carotene, β-carotene, β-cryptoxanthin, lutein, lycopene, and zeaxanthin. All methods demonstrated good performance in the analysis of a composite food standard reference material for the analytes they are applicable to. Methods using two serial connected C18 columns and a mobile phase based on acetonitrile, achieved a better carotenoid separation than the method using a mobile phase based on methanol and one C18-column. Carotenoids from leafy green vegetable matrices appeared to be better extracted with a mixture of methanol and tetrahydrofuran than with tetrahydrofuran alone. Costs of carotenoid determination in foods were lower for the method with mobile phase based on methanol. However for some food matrices and in the case of E-Z isomer separations, this was not technically satisfactory. Food extraction with methanol and tetrahydrofuran with direct evaporation of these solvents, and saponification (when needed) using pyrogallol as antioxidant, combined with a HPLC system using a slight gradient mobile phase based on acetonitrile and a stationary phase composed by two serial connected C18 columns was the most technically and economically favourable method. © 2010.


PubMed | Leiden University, Abel Salazar Biomedical Sciences Institute, Instituto Nacional Of Saude Doutor Ricardo Jorge Insa, Hospital Fernando Fonseca and University of Évora
Type: Journal Article | Journal: PloS one | Year: 2015

Major contributions from pathogen genome analysis and host genetics have equated the possibility of Mycobacterium tuberculosis co-evolution with its human host leading to more stable sympatric host-pathogen relationships. However, the attribution to either sympatric or allopatric categories depends on the resolution or grain of genotypic characterization. We explored the influence on the sympatric host-pathogen relationship of clinical (HIV infection and multidrug-resistant tuberculosis [MDRTB]) and demographic (gender and age) factors in regards to the genotypic grain by using spacer oligonucleotide typing (spoligotyping) for classification of M. tuberculosis strains within the Euro-American lineage. We analyzed a total of 547 tuberculosis (TB) cases, from six year consecutive sampling in a setting with high TB-HIV coinfection (32.0%). Of these, 62.0% were caused by major circulating pathogen genotypes. The sympatric relationship was defined according to spoligotype in comparison to the international spoligotype database SpolDB4. While no significant association with Euro-American lineage was observed with any of the factors analyzed, increasing the resolution with spoligotyping evidenced a significant association of MDRTB with sympatric strains, regardless of the HIV status. Furthermore, distribution curves of the prevalence of sympatric and allopatric TB in relation to patients age showed an accentuation of the relevance of the age of onset in the allopatric relationship, as reflected in the trimodal distribution. On the contrary, sympatric TB was characterized by the tendency towards a typical (standard) distribution curve. Our results suggest that within the Euro-American lineage a greater degree of genotyping fine-tuning is necessary in modeling the biological processes behind the host-pathogen interplay. Furthermore, prevalence distribution of sympatric TB to age was suggestive of host genetic determinisms driven by more common variants.


Andre C.,Instituto Nacional Of Saude Doutor Ricardo Jorge Insa | Jorge F.,New University of Lisbon | Castanheira I.,Instituto Nacional Of Saude Doutor Ricardo Jorge Insa | Matos A.,New University of Lisbon
Journal of Chemometrics | Year: 2013

The development of a chromatographic procedure for an ultra-performance liquid chromatography can be a very time-consuming task, as the general approach for finding the appropriate operating conditions has been a trial-and-error process. The present study reports a novel approach in the field of ultra-performance liquid chromatography by using statistical experimental design based on Taguchi's method, which allows a complete separation of nine isocyanates present in a complex matrix. The resolution between two adjacent peaks was considered as a quality characteristic and transformed to a Taguchi signal-to-noise ratio. An orthogonal array L9 (34) was selected to analyze the effect of four chromatographic factors, that is, proportion of solvent, percent triethylamine (v/v), temperature (°C), and flow (mLmin-1), with three levels each. The joint analysis performed to the significant factors achieved in the eight analyses of variance allowed to identify two methods to conduct a complete separation of all peaks. Six isocyanates were separated with the first method, with all factors at the lowest level. The remaining three isocyanates were separated with the second method, with the proportion of solvent at the highest level and the other factors at the lowest level. The overall Taguchi experimental design identified the proportion of solvent and the flow rate as major chromatographic factors. Finally, confirmatory experiments were performed with samples prepared with six and three isocyanates, confirming the complete separation of all isocyanates in the study. The Taguchi methods provided a systematic and efficient methodology for this optimization, with considerably less effort than would be required for other optimizations techniques. © 2013 John Wiley & Sons, Ltd.


Delgado I.,New University of Lisbon | Andre C.,Instituto Nacional Of Saude Doutor Ricardo Jorge Insa | Ramos A.,New University of Lisbon | Matos A.S.,New University of Lisbon | And 3 more authors.
20th IMEKO World Congress 2012 | Year: 2012

A calibration method was developed for the quantification of isocyanates in cork stoppers agglomerates. Bracketing and multipoint calibration curve was applied to a in-house optimized UPLC/DAD analytical method. The calibration curves were obtained by regression analysis using experimental points that covers the range of concentrations expected in real samples. A contaminated real sample was injected and MDI concentration was 6.2 % and 6.4% when calibration methods to calculate the content were multipoint or bracketing, respectively. According with our results both methods are complementary and necessary to guarantee the reliably of results obtained by chromatographic procedure.


PubMed | Instituto Nacional Of Saude Doutor Ricardo Jorge Insa
Type: Comparative Study | Journal: Journal of chromatography. A | Year: 2010

Three sets of extraction/saponification/HPLC conditions for food carotenoid quantification were technically and economically compared. Samples were analysed for carotenoids alpha-carotene, beta-carotene, beta-cryptoxanthin, lutein, lycopene, and zeaxanthin. All methods demonstrated good performance in the analysis of a composite food standard reference material for the analytes they are applicable to. Methods using two serial connected C(18) columns and a mobile phase based on acetonitrile, achieved a better carotenoid separation than the method using a mobile phase based on methanol and one C(18)-column. Carotenoids from leafy green vegetable matrices appeared to be better extracted with a mixture of methanol and tetrahydrofuran than with tetrahydrofuran alone. Costs of carotenoid determination in foods were lower for the method with mobile phase based on methanol. However for some food matrices and in the case of E-Z isomer separations, this was not technically satisfactory. Food extraction with methanol and tetrahydrofuran with direct evaporation of these solvents, and saponification (when needed) using pyrogallol as antioxidant, combined with a HPLC system using a slight gradient mobile phase based on acetonitrile and a stationary phase composed by two serial connected C(18) columns was the most technically and economically favourable method.

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