FM reconhece financiamento da ANR através de um chaire d&#39;Excellence IDEX Sorbonne Paris Cité, o INSU através de uma subvenção PNP, o Institut Universitaire de France, bem como o programa Labex UniverEarth na Sorbonne Paris Cité (ANR-10-LabX-0023 e ANR -11-IDEX-0005-02). Agradecemos também o financiamento do Conselho Europeu de Investigação no âmbito da Comunidade Europeia H2020 programa-quadro / ERC convenção de subvenção # 637503 (Pristine).

NOTA: Os procedimentos que envolvem animais foram aprovados pelo cuidado e uso Comitê Institucional Animal (IACUC) na Université Paris Diderot. 1. Preparação de Materiais <ol><li> Sub-ferver 1 L de destilar os ácidos (HNO 3, HBr) a fim de purificá-los a partir de impureza. </li><li> Limpe os copos e adaptador de ponta em um quente (~ 100 ° C) HNO 3 banho de ácido concentrado durante pelo menos dois dias. </li><li> Lave as pontas de pipeta em um resfriado 3 N HNO 3 banho por vários dias e enxaguar individualmente três vezes com água deionizada. </li></ol> Preparação 2. Amostra <ol><li> Anestesiar os ratos por injecção intraperitoneal de cetamina e xilazina. Avaliar anestesia pelo método toe pitada. </li><li> Recolhe-se o sangue por punção cardíaca, uma na presença de heparina em tubos de 1,5 ml. </li><li> Separa-se o plasma a partir de células de sangue por centrifugação (10 min, 1500 xg) e transferir o plasma para o polipropileno cfrascos ryogenic usando dicas de polipropileno. </li><li> Remover o sangue restante a partir de órgãos de corte da veia hepática e injectando DPBS através do coração. Avaliar a morte do mouse por deslocamento cervical. </li><li> Colher os órgãos com instrumentos de aço inoxidável estéreis, libertá-los de gordura em torno se houver, e snap-congelá-los em frascos de polipropileno criogênicas. </li></ol> 3. Chemical Purificação <ol><li> Em primeiro lugar, dissolve-se as amostras com uma mistura de ~ 1 ml de concentrado (30%) de H 2 O 2 e ~ 1 ml de concentrado (~ 15 M) HNO3. Será que todos estes passos dentro de um exaustor. <ol><li> Coloque todo o órgão de interesse em uma proveta de 15 ml Teflon. Em seguida, adicione o H 2 O 2 / HNO 3 a 5 o copo. Manter o vaso aberto durante alguns minutos, a fim de evitar salpicos, devido à reacção de oxidação da matéria orgânica e a libertação de CO 2. </li><li> Finalmente, colocar o copo sobre uma placa quente a about 100 ° C durante um par de horas ou até que a solução é perfeitamente claro. </li></ol></li><li> Abrir o recipiente e secar a solução sobre uma placa quente a cerca de 100 ° C. </li><li> Uma vez que a amostra é seca, adicionar 1 ml de HBr 1,5 N para as amostras; fechar a proveta e deixou-se dissolver numa placa quente a 100 ° C durante um par de horas. </li><li> Enquanto isso prepare os 500 colunas ul. <ol><li> Adicionar 500 mL de resina malha AG1X8 200-400 para a coluna e colocá-lo na prateleira coluna com uma taça de lixo abaixo dela. Lavou-se a resina alternando: 5 ml de 18,2 mohms ⋅ cm de água, 5 ml de 0,5 N HNO 3, 5 ml de água, 5 ml de 0,5 N HNO 3, e, em seguida, 5 ml de água. Condiciona-se a resina com 5 ml de HBr 1,5 N. </li></ol></li><li> Retirar os copos da chapa quente e colocá-los em um banho de ultra-som para cerca de 30 min, e em seguida, deixar as taças esfriar a temperatura ambiente. </li><li> Uma vez que o recipiente é arrefecido e a resina é lavada, abrir o recipiente. Coloque o adaptador de ponta para tele seringa, adicione uma ponta de pipeta; Pipete a 1 ml de amostra e carregá-lo sobre a resina (muito lentamente a fim de não agitar a resina). </li><li> Uma vez todo o líquido passa através da coluna, adicionam-se 5 ml de HBr 1,5 N. </li><li> Uma vez que os 5 ml de HBr 1,5 N passar através da coluna, substituir o recipiente de lixo, com uma proveta de 15 ml limpo. </li><li> Adicionar 5 ml de 0,5 N HNO 3 2,5 ml de cada vez. Nesta fase, o Zn é eluído a partir da resina. </li><li> Uma vez 5 ml de HNO 3 passa através da coluna, retirar o copo e colocá-lo sobre uma placa quente a 100 ° C até seca. </li><li> Retirar a coluna do suporte da coluna; lixo a resina (usar uma nova resina para cada amostra). </li><li> Uma vez que a amostra é seca, repetir o protocolo com o mesmo volume de ácidos sobre uma coluna mais pequena (100 ul) e, em seguida, colocá-la numa placa quente até seca. A amostra está agora pronto para espectrometria de massa. </li></ol> 4. espectroscopia de massa de Medição <ol><li> Analisar o compo isotópica Znsição num espectrómetro de massa de plasma indutivamente acoplado-colector multi (MC-ICP-MS). <ol><li> Com os parâmetros de máquina resumidos na Tabela 1. </li></ol></li><li> Posicione os copos de Faraday para coletar a massa (m / z) de 62 Ni, 63 Cu, Zn 64, 65 Cu, Zn 66, 67 Zn e 68 Zn. </li><li> Preparar uma solução contendo 500 ppb Zn em 0,1 M HNO3 para análise isotópica. </li><li> Analisar a solução de 500 ppb de Zn usando uma câmara de pulverização combinada com 100 ul / min nebulizador de teflon. Para cada amostra, medir 30 scans (1 bloco de 30 ciclos), em que o tempo de integração de cada digitalização é 8,389 sec. </li><li> Corrija o fundo subtraindo o zero intensidades sobre-pico a partir de uma solução em branco (o M HNO3 0,1 solução usada para re-dissolver as amostras). </li><li> Controlo e à possível interferência isobárica 64 Ni correto medindo a intensidade do pico de 62 Ni.Suponha que a taxa de 62 64 Ni / Ni é ​​natural (0,2548), este valor correto a partir do viés de massa instrumental, e depois remover o Ni 64 na massa 64 como: 64 Zn reais = 64 Zn medido - 64 = 64 Ni Zn medido - (64 Ni / 62 Ni) naturais x 62 Ni medido. </li><li> Corrigir a polarização massa instrumental por escalonamento cada uma das amostras com uma solução de 500 ppb padrão da norma JMC Lyon Zn (ou outro padrão disponíveis, tais como IMMR-3702). Realizar o escalonamento padrão dividindo a relação de 66 Zn / 64 Zn de amostra pelo valor médio da relação de 66 Zn / 64 Zn de os dois padrões medidos antes e depois da amostra de menos de 1 e multiplicado por 1.000 (ver equação 1). Precisão externo típico no padrão JMC Lyon Zn é de 0,05 permil / amu (2 desvio padrão, 2 sd). </li></ol>

<ol>
	<li>Marechal, C. N., Telouk, P., Albarede, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Precise+analysis+of+copper+and+zinc+isotopic+compositions+by+plasma-source+mass+spectrometry.">Precise analysis of copper and zinc isotopic compositions by plasma-source mass spectrometry.</a> Chemical Geology. 156 (1), 251-273 (1999).</li><li>Marechal, C. N., Albarede, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ion-exchange+fractionation+of+copper+and+zinc+isotopes.">Ion-exchange fractionation of copper and zinc isotopes.</a> Geochim. Cosmochim. Acta. 66 (9), 1499-1509 (2001).</li><li>Weiss, D. J., Mason, T. F. D., Zhao, F. J., Kirk, G. J. D., Coles, B. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isotopic+discrimination+of+zinc+in+higher+plants.">Isotopic discrimination of zinc in higher plants.</a> New Phytologist. 165 (3), 703-710 (2005).</li><li>Jouvin, D., Louvat, P., N, M. F. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Zinc+isotopic+fractionation:+why+organic+matters.">Zinc isotopic fractionation: why organic matters.</a> Environ Sci Technol. 43 (15), 5747-5754 (2009).</li><li>Moynier, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isotopic+fractionation+and+transport+mechanisms+of+Zn+in+plants.">Isotopic fractionation and transport mechanisms of Zn in plants.</a> Chemical Geology. 267 (3-4), 125-130 (2009).</li><li>Moynier, F., Herzog, G., Albarede, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isotopic+composition+of+zinc,+copper,+and+iron+in+lunar+samples.">Isotopic composition of zinc, copper, and iron in lunar samples.</a> Geochim. Cosmochim. Acta. 70 (24), 6103-6117 (2006).</li><li>Moynier, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isotopic+fractionation+of+zinc+in+tektites.">Isotopic fractionation of zinc in tektites.</a> Earth Planet. Sci. Lett. 277 (3-4), 482-489 (2009).</li><li>Chen, H., Savage, P., Teng, F. Z., Helz, R., Moynier, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Zinc+isotope+fractionation+during+magmatic+differentiation+and+the+isotopic+composition+of+the+bulk+Earth.">Zinc isotope fractionation during magmatic differentiation and the isotopic composition of the bulk Earth.</a> Earth Planet. Sci. Lett. 369-370, 34-42 (2013).</li><li>Moeller, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Calibration+of+the+new+certified+materials+ERM-AE633+and+ERM-AE6447+for+copper+and+IRMM+3702+for+zinc+isotope+amount+ratio+determination.">Calibration of the new certified materials ERM-AE633 and ERM-AE6447 for copper and IRMM 3702 for zinc isotope amount ratio determination.</a> Geostd. Geoan. Res. 36 (2), 177-199 (2012).</li><li>Bigeleisen, J., Mayer, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Calculation+of+equilibrium+constants+for+isotopic+exchange+reactions.">Calculation of equilibrium constants for isotopic exchange reactions.</a> J. Chem. Phys. 15, 261-267 (1947).</li><li>Luck, J. M., Ben Othman, D., Albarede, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Zn+and+Cu+isotopic+variations+in+chondrites+and+iron+meteorites:+Early+solar+nebula+reservoirs+and+parent-body+processes.">Zn and Cu isotopic variations in chondrites and iron meteorites: Early solar nebula reservoirs and parent-body processes.</a> Geochim. Cosmochim. Acta. 69 (22), 5351-5363 (2005).</li><li>Moynier, F., Dauphas, N., Podosek, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Search+for+70Zn+Anomalies+in+Meteorites.">A Search for 70Zn Anomalies in Meteorites.</a> Astrophys. J. 700 (2), L92-L95 (2009).</li><li>Paniello, R., Day, J., Moynier, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Zn+isotope+evidence+for+the+origin+of+the+Moon.">Zn isotope evidence for the origin of the Moon.</a> Nature. 490 (7420), 376-380 (2012).</li><li>Pichat, S., Douchet, C., Albarede, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Zinc+isotope+variations+in+deep-sea+carbonates+from+the+eastern+equatorial+Pacific+over+the+last+175+ka.">Zinc isotope variations in deep-sea carbonates from the eastern equatorial Pacific over the last 175 ka.</a> Earth and Planetary Science Letters. 210 (1-2), 167-178 (2003).</li><li>Moynier, F., Fujii, T., Shaw, A., Le Borgne, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Heterogeneous+of+natural+Zn+isotopes+in+mice.">Heterogeneous of natural Zn isotopes in mice.</a> Metallomics. 5 (6), 693-699 (2013).</li><li>Balter, V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bodily+variability+of+zinc+natural+isotope+abundance+in+sheep.">Bodily variability of zinc natural isotope abundance in sheep.</a> Rapid Com. Mass. Spec. 24, 605-612 (2010).</li><li>Balter, V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contrasting+Cu,+Fe,+and+Zn+isotopic+patterns+in+organs+and+body+fluids+of+mice+and+sheep,+with+emphasis+on+cellular+fractionation.">Contrasting Cu, Fe, and Zn isotopic patterns in organs and body fluids of mice and sheep, with emphasis on cellular fractionation.</a> Metallomics. 5 (11), 1470-1482 (2010).</li><li>Urgast, D. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Zinc+isotope+ratio+imaging+of+rat+brain+thin+sections+from+stable+isotope+tracer+by+LA-MC-ICP-MS.">Zinc isotope ratio imaging of rat brain thin sections from stable isotope tracer by LA-MC-ICP-MS.</a> Metallomics. 4, 1057-1063 (2012).</li><li>Marin-Carbonne, J., Rollion-Bard, C., Luais, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In-situ+measurements+of+iron+isotopes+by+SIMS:+MC-ICP-MS+intercalibration+and+application+to+a+magnetite+crystal+from+the+Gunflint+chert.">In-situ measurements of iron isotopes by SIMS: MC-ICP-MS intercalibration and application to a magnetite crystal from the Gunflint chert.</a> Chem. Geol. 285 (1-4), 50-61 (2011).</li><li>Fietzke, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Boron+isotope+ratio+determination+in+carbonates+via+LA-MC-ICP-MS+using+soda-lime+glass+standards+as+reference+material.">Boron isotope ratio determination in carbonates via LA-MC-ICP-MS using soda-lime glass standards as reference material.</a> J. Anal. Atom. Spec. 25, 1953-1957 (2010).</li></ol>

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A reprodutibilidade das medidas é avaliada através de análises replicadas das mesmas amostras realizadas durante as diferentes sessões de análise. Por exemplo 6, que foram replicados da mesma rocha terrestre 7 vezes e obtivemos os resultados reportados na Tabela 2. Como esperado a partir da teoria de fracionamento isotópico 10 e tal como medido em qualquer material sistema solar até agora (por exemplo, meteorito 11-13, plantas...

A medição de alta precisão (melhor do que 100 ppm / unidade de massa atômica) de zinco composição dos isótopos estáveis ​​só foi possível por cerca de 15 anos, graças ao desenvolvimento de multi-coletor de código-fonte plasma massa-espectrómetros e desde então tem sido aplicado principalmente em Terra e ciências planetárias. Os aplicativos para a área médica são novos e têm um forte potencial como biomarcadores para doenças que modificam o metabolismo de zinco (por exemplo, a doença de Alzheimer). Este artigo relata um método para medir com alta precisão as proporções de isótopos estáveis ​​naturais de zinco em vários órgãos do mouse. O mesmo seria aplicável a amostras humanas. O método consiste na dissolução dos órgãos, a purificação química de zinco a partir do resto dos átomos, e, em seguida, a análise da proporção isótopo num espectrómetro de massa. A qualidade de Zn medições isotópicas está dependente da qualidade da purificação química (pureza de Zn, baixo amostra em brancoared para a quantidade de Zn presente na amostra, maior rendimento de produtos químicos do processo) e no controlo da polarização instrumental. O elevado grau de pureza da fracção de Zn final é necessária para remover ambas as interferências isobáricas e não-interferência isobárica que criam um efeito de matriz. Nuclides isobáricas criar interferências diretas (por exemplo, 64 Ni). Interferências não-isobáricas gerar o chamado efeito de &quot;matriz&quot; e alterar a precisão analítica das medições, alterando o estado de ionização por comparação com o padrão de zinco puro a que as amostras são comparados com um. Um baixo em branco (&lt;10 ng) indica que não existe contaminação das amostras por Zn externo que distorceria a composição isotópica medido. Como isótopos de Zn pode ser fraccionado durante a cromatografia de permuta iónica 2, o conjunto de todos os átomos de Zn assegura que não ocorre fraccionamento isotópica, o que implica que o processo químico deve ter um rendimento total. Finalmente, a correção do fracionamento isotópico instrumental durante a medição espectrometria de massa é feita através do método de &quot;bracketing padrão&quot;. Portanto, as principais dificuldades para a obtenção de medições precisas está controlando a contaminação externa (ou seja, baixo em branco), produzindo uma purificação química rendimento total que é limpo de quaisquer outros átomos ou moléculas, e corrigir o fracionamento isotópico instrumental na massa espectrômetro. Neste artigo vamos descrever o nosso protocolo analítico para separar Zn dos órgãos do rato, bem como as medidas de espectrometria de massa. A extração é feita usando uma baixa quantidade de ácidos diluídos (HBr / HNO3 media) em micro-colunas (0,5 ul e 0,1 ul) de resina de troca aniônica. Tem um rendimento total e as medições tem uma reprodutibilidade externa melhor do que 50 ppm em relação a 66 Zn / 64 Zn. Outra vantagem da metanfetaminaOD é que ele é muito rápido. O método é, portanto, muito bem adaptado para ciências médicas, em que é necessário analisar um grande número de amostras em comparação com geosciences, onde estes foram desenvolvidos métodos analíticos.

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			<td height="42" style="height:42px;width:236px;">Multi-collection inductively-coupled-plasma mass-spectromter</td>
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			<td colspan="2">140-1443-MSDS</td>
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			<td height="21" style="height:21px;width:236px;">teflon beakers</td>
			<td style="width:99px;">Savillex&#160;</td>
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			<td height="42" style="height:42px;width:236px;">Home-made teflon colunms made with shrinkable teflon</td>
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high precision zinc isotopic measurements applied to mouse organs

Nós apresentamos um procedimento para medir com alta precisão as taxas de isótopos de zinco em órgãos do mouse. O zinco é composto por 5 isótopos estáveis ​​(64 Zn, 66 Zn, 67 Zn 68, Zn e Zn 70), que são naturalmente fraccionados entre órgãos do mouse. Nós primeiro mostrar como dissolver os diferentes órgãos, a fim de libertar os átomos de Zn; este passo é realizado por uma mistura de HNO 3 e H 2 O 2. Em seguida, purificar os átomos de zinco a partir de todos os outros elementos, em particular de interferências isobáricas (por exemplo, NI), por cromatografia de troca aniônica em um diluído HBr / HNO meio 3. Estas duas primeiras etapas são realizadas em um laboratório limpo usando produtos químicos de alta pureza. Finalmente, as proporções de isótopos são medidos utilizando um multi-colector espectrómetro de massa acoplado indutivamente de plasma, em baixa resolução. As amostras são injectados usando uma câmara de pulverização e o fraccionamento isotópica induzida pelo espectrómetro de massa é o corrected, comparando o rácio das amostras para o rácio de um padrão (técnica de escalonamento padrão). Este procedimento típico completo produz uma razão isotópica com uma reprodutibilidade 50 ppm (2 sd).

Em 1,5 N HBr, as principais espécies de zinco (ZnBr3-) forma complexos muito fortes com a resina de troca aniônica, enquanto a maioria dos outros elementos não interagem com a resina. O zinco é, então, recuperado, alterando a forma diluída de HNO 3, mudando a especiação de Zn para o Zn2 + que é libertado a partir da resina de 6,7. As taxas de isótopos são tipicamente expressos em partes por mil desvios relativos a um padrão: <p class="jove_con...

Watch this Scientific Journal Video about High Precision Zinc Isotopic Measurements Applied to Mouse Organs at JoVE.com

High Precision Zinc Isotopic Measurements Applied to Mouse Organs

We present the technique to measure with high precision zinc isotope ratios in mouse organs.

Medições de alta precisão de zinco isotópicas Aplicadas à mouse Órgãos

We present a procedure to measure with high precision zinc isotope ratios in mouse organs. Zinc is composed of 5 stable isotopes (64Zn, 66Zn, 67Zn, 68Zn ...

high-precision-zinc-isotopic-measurements-applied-to-mouse-organs

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Chemistry

7.4K visualizações.  Université Paris Diderot. We present the technique to measure with high precision zinc isotope ratios in mouse organs.

Vídeo: High Precision Zinc Isotopic Measurements Applied to Mouse Organs

High-pressure Sapphire Cell for Phase Equilibria Measurements of CO2/Organic/Water Systems

The high pressure sapphire cell apparatus was constructed to visually determine the composition of multiphase systems without physical sampling. Specifically, the sapphire cell enables visual data collection from multiple loadings to solve a set of material balances to precisely determine phase composition. Ternary phase diagrams can then be established to determine the proportion of each component in each phase at a given condition. In principle, any ternary system can be studied although ternary systems (gas-liquid-liquid) are the specific examples discussed herein. For instance, the ternary THF-Water-CO2 system was studied at 25 and 40&#160;&#176;C and is described herein. Of key importance, this technique does not require sampling. Circumventing the possible disturbance of the system equilibrium upon sampling, inherent measurement errors, and technical difficulties of physically sampling under pressure is a significant benefit of this technique. Perhaps as important, the sapphire cell also enables the direct visual observation of the phase behavior. In fact, as the CO2 pressure is increased, the homogeneous THF-Water solution phase splits at about 2 MPa. With this technique, it was possible to easily and clearly observe the cloud point and determine the composition of the newly formed phases as a function of pressure.
The data acquired with the sapphire cell technique can be used for many applications. In our case, we measured swelling and composition for tunable solvents, like gas-expanded liquids, gas-expanded ionic liquids and Organic Aqueous Tunable Systems (OATS)1-4. For the latest system, OATS, the high-pressure sapphire cell enabled the study of (1) phase behavior as a function of pressure and temperature, (2) composition of each phase (gas-liquid-liquid) as a function of pressure and temperature and (3) catalyst partitioning in the two liquid phases as a function of pressure and composition. Finally, the sapphire cell is an especially effective tool to gather accurate and reproducible measurements in a timely fashion.

High-pressure Sapphire Cell for Phase Equilibria Measurements of CO2/Organic/Water Systems

The high pressure sapphire cell apparatus is a unique tool to study, without sampling, phase behavior under a wide range of pressures. Using a cathetometer, very precise volume measurements can be recorded to measure liquid expansion and phase composition. Thus, this synthetic method enables the study of (1) phase equilibria of multi-component mixtures and (2) the partition behavior of catalyst or model compounds as a function of pressure.

Sapphire celular de alta pressão para medições de Equilíbrio de Fases de CO<sub&gt; 2</sub&gt; / Orgânicos / Sistemas de Água

The high pressure sapphire cell apparatus was constructed to visually determine the composition of multiphase systems without physical sampling. ...

Química

Cooling Rate Dependent Ellipsometry Measurements to Determine the Dynamics of Thin Glassy Films

This report aims to fully describe the experimental technique of using ellipsometry for cooling rate dependent Tg (CR-Tg) experiments. These measurements are simple high-throughput characterization experiments, which can determine the glass transition temperature (Tg), average dynamics, fragility and the expansion coefficient of the super-cooled liquid and glassy states for a variety of glassy materials. This technique allows for these parameters to be measured in a single experiment, while other methods must combine a variety of different techniques to investigate all of these properties. Measurements of dynamics close to Tg are particularly challenging. The advantage of cooling rate dependent Tg measurements over other methods which directly probe bulk and surface relaxation dynamics is that they are relatively quick and simple experiments, which do not utilize fluorophores or other complicated experimental techniques. Furthermore, this technique probes the average dynamics of technologically relevant thin films in temperature and relaxation time (&#964;&#945;) regimes relevant to the glass transition (&#964;&#945; &#62; 100 sec). The limitation to using ellipsometry for cooling rate dependent Tg experiments is that it cannot probe relaxation times relevant to measurements of viscosity (&#964;&#945; &#60;&#60; 1 sec). Other cooling rate dependent Tg measurement techniques, however, can extend the CR-Tg method to faster relaxation times. Furthermore, this technique can be used for any glassy system so long as the integrity of the film remains throughout the experiment.

Here, we present a protocol for cooling rate dependent ellipsometry experiments, which can determine the glass transition temperature (Tg), average dynamics, fragility and the expansion coefficient of the super-cooled liquid and glass for a variety of glassy materials.

Arrefecimento Medidas de freqüência Elipsometria Dependentes para determinar a dinâmica de finas Glassy Films

This report aims to fully describe the experimental technique of using ellipsometry for cooling rate dependent Tg (CR-Tg) experiments. These measurements ...

Immunostaining Phospho-epitopes in Ciliated Organs of Whole Mount Zebrafish Embryos

The rapid proliferation of cells, the tissue-specific expression of genes and the emergence of signaling networks characterize early embryonic development of all vertebrates. The kinetics and location of signals - even within single cells - in the developing embryo complements the identification of important developmental genes. Immunostaining techniques are described that have been shown to define the kinetics of intracellular and whole animal signals in structures as small as primary cilia. The techniques for fixing, imaging and processing images using a laser-scanning confocal compound microscope can be completed in as few as 36 hr.
Zebrafish (Danio rerio) is a desirable organism for investigators who seek to conduct studies in a vertebrate species that is affordable and relevant to human disease. Genetic knockouts or knockdowns must be confirmed by the loss of the actual protein product. Such confirmation of protein loss can be achieved using the techniques described here. Clues into signaling pathways can also be deciphered by using antibodies that are reactive with proteins that have been post-translationally modified by phosphorylation. Preserving and optimizing the phosphorylated state of an epitope is therefore critical to this determination and is accomplished by this protocol.
This study describes techniques to fix embryos during the first 72 hr of development and co-localize a variety of relevant epitopes with cilia in the Kupffer&#39;s Vesicle (KV), the kidney and the inner ear. These techniques are straightforward, do not require dissection and can be completed in a relatively short period of time. Projecting confocal image stacks into a single image is a useful means of presenting these data.

Techniques are described to immunostain phospho-epitopes in whole zebrafish embryos and then conduct two-color fluorescent confocal localization in cellular structures as small as primary cilia. The techniques for fixing and imaging can define the location and kinetics of the appearance or activation of specific proteins.

Imunocoloração fosfo-epitopos em ciliadas Órgãos da Whole Mount embriões Zebrafish

The rapid proliferation of cells, the tissue-specific expression of genes and the emergence of signaling networks characterize early embryonic development ...

Characterization, Quantification and Compound-specific Isotopic Analysis of Pyrogenic Carbon Using Benzene Polycarboxylic Acids (BPCA)

Fire-derived, pyrogenic carbon (PyC), sometimes called black carbon (BC), is the carbonaceous solid residue of biomass and fossil fuel combustion, such as char and soot. PyC is ubiquitous in the environment due to its long persistence, and its abundance might even increase with the projected increase in global wildfire activity and the continued burning of fossil fuel. PyC is also increasingly produced from the industrial pyrolysis of organic wastes, which yields charred soil amendments (biochar). Moreover, the emergence of nanotechnology may also result in the release of PyC-like compounds to the environment. It is thus a high priority to reliably detect, characterize and quantify these charred materials in order to investigate their environmental properties and to understand their role in the carbon cycle.
Here, we present the benzene polycarboxylic acid (BPCA) method, which allows the simultaneous assessment of PyC&#39;s characteristics, quantity and isotopic composition (13C and 14C) on a molecular level. The method is applicable to a very wide range of environmental sample materials and detects PyC over a broad range of the combustion continuum, i.e., it is sensitive to slightly charred biomass as well as high temperature chars and soot. The BPCA protocol presented here is simple to employ, highly reproducible, as well as easily extendable and modifiable to specific requirements. It thus provides a versatile tool for the investigation of PyC in various disciplines, ranging from archeology and environmental forensics to biochar and carbon cycling research.

Characterization, Quantification and Compound-specific Isotopic Analysis of Pyrogenic Carbon Using Benzene Polycarboxylic Acids BPCA

We present the benzene polycarboxylic acid (BPCA) method for assessing pyrogenic carbon (PyC) in the environment. The compound-specific approach uniquely provides simultaneous information about the characteristics, quantity and isotopic composition (13C and 14C) of PyC.

Caracterização, quantificação e específicos do Composto isotópica Análise de Pirogênica carbono, usando Benzeno policarboxílicos Acids (BPCA)

Fire-derived, pyrogenic carbon (PyC), sometimes called black carbon (BC), is the carbonaceous solid residue of biomass and fossil fuel combustion, such as ...

Fizzy Extraction of Volatile Organic Compounds Combined with Atmospheric Pressure Chemical Ionization Quadrupole Mass Spectrometry

Chemical analysis of volatile and semivolatile compounds dissolved in liquid samples can be challenging. The dissolved components need to be brought to the gas phase, and efficiently transferred to a detection system. Fizzy extraction takes advantage of the effervescence phenomenon. First, a carrier gas (here, carbon dioxide) is dissolved in the sample by applying overpressure and stirring the sample. Second, the sample chamber is decompressed abruptly. Decompression leads to the formation of numerous carrier gas bubbles in the sample liquid. These bubbles assist the release of the dissolved analyte species from the liquid to the gas phase. The released analytes are immediately transferred to the atmospheric pressure chemical ionization interface of a triple quadrupole mass spectrometer. The ionizable analyte species give rise to mass spectrometric signals in the time domain. Because the release of the analyte species occurs over short periods of time (a few seconds), the temporal signals have high amplitudes and high signal-to-noise ratios. The amplitudes and areas of the temporal peaks can then be correlated with concentrations of the analytes in the liquid samples subjected to fizzy extraction, which enables quantitative analysis. The advantages of fizzy extraction include: simplicity, speed, and limited use of chemicals (solvents).

Fizzy extraction is a new laboratory technique for analysis of volatile and semivolatile compounds. A carrier gas is dissolved in the liquid sample by applying overpressure and stirring the sample. The sample chamber is then decompressed. The analyte species are liberated to the gas phase due to effervescence.

Extração efervescente de compostos orgânicos voláteis combinados com a espectrometria de massa de quadrupoolos de ionização química de pressão atmosférica

Chemical analysis of volatile and semivolatile compounds dissolved in liquid samples can be challenging. The dissolved components need to be brought to ...

Aerosol-assisted Chemical Vapor Deposition of Metal Oxide Structures: Zinc Oxide Rods

Whilst columnar zinc oxide (ZnO) structures in the form of rods or wires have been synthesized previously by different liquid- or vapor-phase routes, their high cost production and/or incompatibility with microfabrication technologies, due to the use of pre-deposited catalyst-seeds and/or high processing temperatures exceeding 900 &#176;C, represent a drawback for a widespread use of these methods. Here, however, we report the synthesis of ZnO rods via a non-catalyzed vapor-solid mechanism enabled by using an aerosol-assisted chemical vapor deposition (CVD) method at 400 &#176;C with zinc chloride (ZnCl2) as the precursor and ethanol as the carrier solvent. This method provides both single-step formation of ZnO rods and the possibility of their direct integration with various substrate types, including silicon, silicon-based micromachined platforms, quartz, or high heat resistant polymers. This potentially facilitates the use of this method at a large-scale, due to its compatibility with state-of-the-art microfabrication processes for device manufacture. This report also describes the properties of these structures (e.g., morphology, crystalline phase, optical band gap, chemical composition, electrical resistance) and validates its gas sensing functionality towards carbon monoxide.

Columnar zinc oxide structures in the form of rods are synthesized via aerosol-assisted chemical vapor deposition without the use of pre-deposited catalyst-seed particles. This method is scalable and compatible with various substrates based on either silicon, quartz or polymers.

Aerossol-assistida de deposição de Vapor químico de estruturas de óxido de Metal: hastes de óxido de zinco

Whilst columnar zinc oxide (ZnO) structures in the form of rods or wires have been synthesized previously by different liquid- or vapor-phase routes, ...

High Resolution Physical Characterization of Single Metallic Nanoparticles

Individual molecules can be detected and characterized by measuring the degree by which they reduce the ionic current flowing through a single nanometer-scale pore. The signal is characteristic of the molecule&#39;s physicochemical properties and its interactions with the pore. We demonstrate that the nanopore formed by the bacterial protein exotoxin Staphylococcus aureus alpha hemolysin (&#945;HL) can detect polyoxometalates (POMs, anionic metal oxygen clusters), at the single molecule limit. Moreover, multiple degradation products of 12-phosphotungstic acid POM (PTA, H3PW12O40) in solution are simultaneously measured. The single molecule sensitivity of the nanopore method allows for POMs to be characterized at significantly lower concentrations than required for nuclear magnetic resonance (NMR) spectroscopy. This technique could serve as a new tool for chemists to study the molecular properties of polyoxometalates or other metallic clusters, to better understand POM synthetic processes, and possibly improve their yield. Hypothetically, the location of a given atom, or the rotation of a fragment in the molecule, and the metal oxidation state could be investigated with this method. In addition, this new technique has the advantage of allowing the real-time monitoring of molecules in solution.&#160;

Here, we present a protocol to detect discrete metal oxygen clusters, polyoxometalates (POMs), at the single molecule limit using a biological nanopore-based electronic platform. The method provides a complementary approach to traditional analytical chemistry tools used in the study of these molecules.

Caracterização física de alta resolução de nanopartículas metálicas únicas

Individual molecules can be detected and characterized by measuring the degree by which they reduce the ionic current flowing through a single ...

Isotopic Effect in Double Proton Transfer Process of Porphycene Investigated by Enhanced QM/MM Method

The single deuterium substitution in porphycene leads to an asymmetric molecular geometry, which may affect the double proton transfer process in the porphycene molecule. In this study, we applied an enhanced QM/MM method called SITS-QM/MM to investigate hydrogen/deuterium (H/D) isotope effects on the double proton transfer in porphycene. Distance changes in SITS-QM/MM molecular dynamics simulations suggested that the deuterium substituted porphycene adopted the stepwise double proton transfer mechanism. The structural analysis and the free energy shifts of double proton transfer process indicated that the asymmetric isotopic substitution subtly compressed the covalent hydrogen bonds and may alter the original transition state location.

A protocol that uses enhanced QM/MM method to investigate the isotopic effect on the double proton transfer process in porphycene is presented here.

Efeito isotópico no processo de transferência de prótons duplo de Porficeno investigado por método Enhanced QM/MM

The single deuterium substitution in porphycene leads to an asymmetric molecular geometry, which may affect the double proton transfer process in the ...

Ultrafast Time-resolved Near-IR Stimulated Raman Measurements of Functional &#960;-conjugate Systems

Femtosecond time-resolved stimulated Raman spectroscopy is a promising method of observing the structural dynamics of short-lived transients with near infrared (near-IR) transitions, because it can overcome the low sensitivity of spontaneous Raman spectrometers in the near-IR region. Here, we describe technical details of a femtosecond time-resolved near-IR multiplex stimulated Raman spectrometer that we have recently developed. A description of signal generation and optimization, measurement, data acquisition, and calibration and correction of recorded data is provided as well. We present an application of our spectrometer to analyze the excited-state dynamics of &#946;-carotene in toluene solution. A C=C stretch band of &#946;-carotene in the second lowest excited singlet (S2) state and the lowest excited singlet (S1) state is clearly observed in the recorded time-resolved stimulated Raman spectra. The femtosecond time-resolved near-IR stimulated Raman spectrometer is applicable to the structural dynamics of &#960;-conjugate systems from simple molecules to complex materials.

Details of signal generation and optimization, measurement, data acquisition, and data handling for a femtosecond time-resolved near-IR stimulated Raman spectrometer are described. A near infrared stimulated Raman study on the excited-state dynamics of &#946;-carotene in toluene is shown as a representative application.

Mediçãos de Raman estimuladas do tempo ultrarrápido de sistemas funcionais de πconjugados

Femtosecond time-resolved stimulated Raman spectroscopy is a promising method of observing the structural dynamics of short-lived transients with near ...

Sample Preparation in Quartz Crystal Microbalance Measurements of Protein Adsorption and Polymer Mechanics

In this study, we present various examples of how thin film preparation for quartz crystal microbalance experiments informs the appropriate modeling of the data and determines which properties of the film can be quantified. The quartz crystal microbalance offers a uniquely sensitive platform for measuring fine changes in mass and/or mechanical properties of an applied film by observing the changes in mechanical resonance of a quartz crystal oscillating at high frequency. The advantages of this approach include its experimental versatility, ability to study changes in properties over a wide range of experimental time lengths, and the use of small sample sizes. We demonstrate that, based on the thickness and shear modulus of the layer deposited on the sensor, we can acquire different information from the material. Here, this concept is specifically exploited to display experimental parameters resulting in mass and viscoelastic calculations of adsorbed collagen on gold and polyelectrolyte complexes during swelling as a function of salt concentration.

The quartz crystal microbalance can provide accurate mass and viscoelastic properties for films in the micron or submicron range, which is relevant for investigations in biomedical and environmental sensing, coatings, and polymer science. The sample thickness influences which information can be obtained from the material in contact with the sensor.

Preparação de amostras em medições de microequilíbrio de cristal de quartzo de adsorção de proteínas e mecânica de polímeros

In this study, we present various examples of how thin film preparation for quartz crystal microbalance experiments informs the appropriate modeling of ...