Name: nmr spectroscopy as a robust tool for the rapid evaluation of the lipid profile of fish oil supplements
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Description: Watch this Scientific Journal Video about NMR Spectroscopy as a Robust Tool for the Rapid Evaluation of the Lipid Profile of Fish Oil Supplements at JoVE.com

Este trabajo fue apoyado por los alimentos para la salud descubrimiento temático en la Universidad Estatal de Ohio y el Departamento de Ciencia y Tecnología de Alimentos de la Universidad Estatal de Ohio. Los autores desean agradecer a las instalaciones de RMN en la Universidad Estatal de Ohio y la facilidad de RMN en la Universidad Estatal de Pensilvania.

1. Preparación de la muestra de RMN Nota: Precaución, consulte todas las hojas de datos de seguridad (MSDS) antes de usar. Cloroformo deuterado (CDCl3) usado en la preparación de muestras es tóxico. Por favor, use todas las prácticas apropiadas de seguridad al realizar la preparación de muestras que incluye el uso de una campana de humos y equipo de protección personal (gafas de seguridad, guantes, bata de laboratorio, pantalones largos, cerrado-dedo del pie zapatos). <ol...

<ol>
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B., 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=Standardless+1H+NMR+determination+of+pharmacologically+active+substances+in+dietary+supplements+and+medicines+that+have+been+illegally+traded+over+the+internet.">Standardless 1H NMR determination of pharmacologically active substances in dietary supplements and medicines that have been illegally traded over the internet.</a> Drug Test. Anal. 5 (6), 400-411 (2013).</li><li>Berger, S., Braun, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> 200 and more NMR experiments: a practical course. , (2004).</li><li>Knothe, G., Kenar, J. 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Food Chem. 41 (8), 1247-1253 (1993).</li><li>Dais, P., Misiak, M., Hatzakis, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Analysis+of+marine+dietary+supplements+using+NMR+spectroscopy.">Analysis of marine dietary supplements using NMR spectroscopy.</a> Anal. Methods. 7 (12), 5226-5238 (2015).</li><li>Pickova, J., Dutta, P. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cholesterol+Oxidation+in+Some+Processed+Fish+Products.">Cholesterol Oxidation in Some Processed Fish Products.</a> J. Anal. Oil Chem. Soc. 80 (10), 993-996 (2003).</li><li>Siddiqui, N., Sim, J., Silwood, C. J. L., Toms, H., Iles, R. 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Modificaciones y estrategias para la solución de problemas Calidad espectral. La anchura de línea de la señal de RMN y por lo tanto la resolución del espectro de RMN depende de cuñas, que es un proceso para la optimización de la homogeneidad del campo magnético altamente. Para el análisis de rutina, 1D calce es adecuada y no se requiere un calce 3D, dado que se lleva a cabo por el personal de RMN sobre una base regular. Si este no es el caso, un calce 3D debe realizarse...

El consumo de n -3 ácidos grasos en la dieta ha demostrado ser beneficioso contra varias condiciones tales como trastornos del corazón 1, 2, 3, enfermedades inflamatorias 4 y diabetes 5. La dieta occidental es considerada pobre en ácidos grasos n -3 y por lo tanto se recomienda el consumo de suplementos de aceite de pescado para mejorar la n -6 / n -3 equilibrio en la nutrición de los consumidores 1. A pesar del reciente aumento en el consumo de suplemento de aceite de pescado, sigue habiendo dudas sobre la seguridad, la autenticidad y la calidad de algunos de estos productos. El análisis de la composición rápida y precisa de los suplementos de aceite de pescado es esencial para evaluar adecuadamente la calidad de estos productos comerciales y garantizar la seguridad del consumidor. Las metodologías más comunes para la evaluación de suplemento de aceite de pescados son la cromatografía de gases (GC) y espectroscopia infrarroja (IR). Si bien estos son métodos altamente sensibles, sufren de varios inconvenientes 6. El análisis de GC es que consume tiempo (4-8 h), ya que la separación y la derivatización de compuestos individuales se requiere 7 y la oxidación de lípidos se pueden producir durante el análisis 8, 9. Aunque la espectroscopia de IR puede ser cuantitativa, se requiere un modelo de predicción a ser construido usando regresión de mínimos cuadrados parciales (PLSR), aunque hay excepciones en el que bandas de IR puede ser atribuida a un solo compuesto 10. PLSR requiere el análisis de un gran número de muestras, lo que aumenta el tiempo del análisis 11. Por esta razón, hay un interés creciente en el desarrollo de nuevas metodologías analíticas que permiten un análisis preciso y rápido de un gran número de muestras de aceite de pescado. Organizaciones como la OfiOficina de Suplementos Dietéticos (ODS) de los Institutos Nacionales de Salud (NIH) y la Administración de Alimentos y Medicamentos (FDA) han colaborado con la Asociación de Químicos Analíticos Oficiales (AOAC) para desarrollar estos nuevos métodos 12, 13. Uno de los métodos analíticos más prometedores para la detección y la evaluación de las matrices de múltiples componentes, tales como suplementos dietéticos, es la espectroscopia de resonancia magnética nuclear (RMN) 14, 15. espectroscopía de RMN tiene varias ventajas: es una técnica no destructiva y cuantitativa, se requiere un mínimo o ningún preparación de la muestra, y se caracteriza por una excelente precisión y reproducibilidad. Además, la espectroscopía de RMN es una metodología respetuosa del medio ambiente, ya que utiliza sólo pequeñas cantidades de disolventes. El principal inconveniente de la espectroscopía de RMN es su sensibilidad relativamente baja en comparación con otros analytimétodos Cal, sin embargo, los recientes avances tecnológicos en la instrumentación, como los campos más fuertes magnéticos, sondas criogénicas de varios diámetros, procesamiento de datos avanzado, y secuencias de pulsos versátiles y técnicas han aumentado la sensibilidad hasta el rango de nM. Mientras RMN instrumentación es de alto costo, el tiempo de vida de los espectrómetros de RMN y las muchas aplicaciones de RMN reducir el costo de los análisis en el largo plazo. Este protocolo de vídeo detallada está destinada a ayudar a los nuevos practicantes en el campo evitar los errores asociados con la 1 H y 13 C RMN análisis espectroscópico de suplementos de aceite de pescado.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Avance III 850 NMR instrument</td>
			<td>Bruker</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Avance III 500 NMR instrument</td>
			<td>Bruker</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>TCI 5mm probe</td>
			<td>Bruker</td>
			<td></td>
			<td>Helium cooled inverse (proton deetected) NMR probe featuring three independent channels (1H, 13C, 15N)</td>
		</tr>
		<tr>
			<td>BBO prodigy 5mm probe</td>
			<td>Bruker</td>
			<td></td>
			<td>Nitrogen cooled observe (X-nuclei detected) probe, featuring two channels; one for 1H and 19F detectionand one for X-nuclei (covering from 15N to 31P)</td>
		</tr>
		<tr>
			<td>Spinner turbin</td>
			<td>Bruker</td>
			<td></td>
			<td>NMR spinners are made by polymer materials and they have a rubber o-ring to hold the NMR tube securely in place</td>
		</tr>
		<tr>
			<td>Topspin 3.5</td>
			<td>Bruker</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>deuterated chloroform</td>
			<td>Sigma-Aldrich</td>
			<td>&#160;865-49-6</td>
			<td>99.8 atom % D, contains 0.03 TMS</td>
		</tr>
		<tr>
			<td>2,6-Di-tert-butyl-4-methylphenol,</td>
			<td>Sigma-Aldrich</td>
			<td>&#160;128-37-0</td>
			<td>purity &#62;99%</td>
		</tr>
		<tr>
			<td>Fish oil samples</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>NMR tubes</td>
			<td>New Era</td>
			<td>NE-RG5-7</td>
			<td>5mm OD Routine &#8220;R&#8221; Series NMR Sample Tube</td>
		</tr>
		<tr>
			<td>BSMS</td>
			<td>Bruker</td>
			<td></td>
			<td>Bruker Systems Management System; control system device</td>
		</tr>
	</tbody>
</table>

nmr spectroscopy as a robust tool for the rapid evaluation of the lipid profile of fish oil supplements

La dieta occidental es deficiente en n -3 ácidos grasos, por lo tanto, se recomienda el consumo de suplementos de aceite de pescado para aumentar la ingesta de estos nutrientes esenciales. El objetivo de este trabajo es demostrar el análisis cualitativo y cuantitativo de los suplementos de aceite de pescado encapsuladas usando de alta resolución 1 H y 13 C espectroscopía de RMN utilizando dos instrumentos de RMN diferentes; un 500 MHz y un instrumento de 850 MHz. Tanto de protones (1 H) y carbono (13 C) Los espectros de RMN se pueden utilizar para la determinación cuantitativa de los constituyentes principales de los suplementos de aceite de pescado. Cuantificación de los lípidos en los suplementos de aceite de pescado se logra mediante la integración de las señales de RMN correspondientes en los espectros 1D relevante. Los resultados obtenidos por 1 H y 13 C RMN están en buen acuerdo con el uno al otro, a pesar de la diferencia en la resolución y la sensibilidad entre los dos núcleos y los dos instrumentos. Oferta 1 H RMNsa análisis más rápido en comparación con el 13 C RMN, como el espectro se puede grabar en menos de 1 min, en contraste con 13 análisis de RMN de C, que dura de 10 min a una hora. El espectro de RMN 13 C, sin embargo, es mucho más informativo. Puede proporcionar datos cuantitativos para un mayor número de ácidos grasos individuales y se puede utilizar para determinar la distribución posicional de los ácidos grasos en el esqueleto de glicerol. Ambos núcleos pueden proporcionar información cuantitativa en un solo experimento sin la necesidad de purificación o separación pasos. La fuerza del campo magnético afecta principalmente a los espectros de RMN 1 H debido a su menor resolución con respecto a 13 C RMN, sin embargo, los instrumentos de RMN incluso menor costo se pueden aplicar eficientemente como un método estándar por los laboratorios de la industria alimentaria y de control de calidad.

1 H y 13 espectros de RMN C se recogieron para los suplementos de aceite de pescado comercialmente disponibles utilizando dos instrumentos de RMN; un 850 MHz y un espectrómetro de 500 MHz. Estos espectros se pueden utilizar para la determinación cuantitativa de los componentes del aceite de pescado, como el ácido docosahexaenoico (DHA) y ácido eicosapentaenoico (EPA), así como otros compuestos tales como n -1 cadenas de acilo y el índice importante nu...

Watch this Scientific Journal Video about NMR Spectroscopy as a Robust Tool for the Rapid Evaluation of the Lipid Profile of Fish Oil Supplements at JoVE.com

NMR Spectroscopy as a Robust Tool for the Rapid Evaluation of the Lipid Profile of Fish Oil Supplements

Aquí, de alta resolución 1 H y 13 C de Resonancia Magnética Nuclear (RMN) se utilizó como una herramienta rápida y fiable para el análisis cuantitativo y cualitativo de suplementos de aceite de pescado encapsuladas.

Espectroscopía de RMN como una herramienta robusta para la Evaluación Rápida del perfil lipídico de los suplementos de aceite de pescado

The western diet is poor in n-3 fatty acids, therefore the consumption of fish oil supplements is recommended to increase the intake of these essential ...

The overall goal of this method is to determine the lipid profile of fish oil supplements using Nuclear Magnetic Resonance spectroscopy. This method can help answer key questions for lipid analysis such as the lipid contents of fish oil supplements and the positional distribution of various fatty acids on the glycerol skeleton. The main advantage of this technique is that it makes the rapid and robust analysis of several different molecules feasible in one snapshot without using separation and purification steps.To prepare the samples for proton and carbon NMR analysis, first extract 120 microliters, which is around 110 milligrams of fish oil, from a dietary capsule using a syringe and place it in a 4-mL glass vial. Record the weight of the fish oil. Use 500 microliters of a BHT stock solution to dissolve 100 milligrams plus or minus 10 milligrams of fish oil.After dissolving the oil, transfer all of the solution directly into a high quality, five millimeter NMR tube and attach a cap. Analyze the samples within 24 hours of preparing the samples. Insert the NMR tube into a spinner turbine.Place the spinner and the tube on the top of a graded depth gauge and gently push the top of the tube until its bottom part touches the bottom of the gauge. Then place the NMR tube in an open spot of the sample case. Note the slot number the sample is placed in.To load the sample in the NMR instrument, return to the control computer and type SX-number, where number is the slot in the sample case holding the sample. Wait for the deuterium signal of deuterated chloroform to appear on the screen. As soon as the deuterium signal is visible, type lock"on the command line and select deuterated chloroform from the solvents list to lock the sample using its deuterium resonance.Type bsmsdisp"in the command line to ensure spinning is not active. If the spin button is green, click it to deactivate spinning. After typing the command new"to create a new data set, enter a name for the data set in the name tab and the experiment number in the experiment number tab titled EXPNO.Use number one in the process number tab titled PROCNO. Write the title of the experiment in the TITLE tab. In the Experiment tab, hit Select and choose the C13IG parameter file.Then hit Set selected item in editor"and click OK.Type getprosol"in the command line to obtain the standard parameters for the current NMR probe and solvent. Then type the command atma"to perform automatic tuning and to matching of the probe for both carbon and proton nuclei. Next, perform one-dimensional gradient shimming to achieve a highly homogeneous magnetic field and thus optimum line shape for the NMR signals.To do so, use the standard automatic procedure for 1D shimming simply by sequently executing the commands qu topshim 1dfast ss;qu topshim tuneb ss;and qu topshim report"in the command line. After parameter optimization as described in the text protocol, acquire one dimensional carbon NMR spectra. To do so, go to the carbon data set and use an inverse gated decoupled pulse sequence by typing pulprog zgig"in the command line.Type commands on the command line to set up the spectral with nppm, the center of the RF transmitter, the number of scans, the number of dummy scans, the number of data points, and the pulse duration for a ninety degree pulse angle. Type digmod baseopt"in the command line to acquire a spectrum with an improved base line. Then set a relaxation delay of 60 seconds for the 850 megahertz instrument by typing di 60s"in the command line.Set the receiver gain to an appropriate value using the command rga"for automatic calculation of receiver gain. Finally, start the acquisition by typing the pulse acquire command zg"in the command line. To process the data, first type si 64K"in the command line to apply zero filling and set the size of the rail spectrum to 64K.Then set the line broadening parameter to 1.0 hertz by typing lb 1.0"in the command line to apply a waiting function with a line broadening factor of 1.0 hertz prior to 4ea transform. Execute 4ea transformation by typing efp"in the command line. Next perform automatic phase correction by typing the command apk"in the command line.If additional phase adjustments are required to further improve the spectrum, click on the Process tab. Then click on the Adjust Phase icon and the Phase Correction icons for zero order and first order phase correction. While clicking on the zero order and first order phase correction icons, drag the mouse until all of the signals are in positive absorption mode.Apply and save the phase correction values by clicking the Return and Save button to exit the Phase Correction mode. Apply a polynomial forth order function for baseline correction upon integration by typing the command abs n"in the command line. To report chemical shifts in ppm from TMS, zoom in on the TMS signal, then click on the Calibration icon and place the cursor with the red line on top of the NMR signal to be referenced, TMS in this case.To adjust the threshold intensity, use the middle mouse button if needed. Left click and type in 0. When using BHT as an internal standard, integrate the peak at delta 151.45 extending five hertz from each side of the peak.Then set the integral equal to the micromoles of BHT per 0.5 milliliters of the stock solution. Integrate the peaks of interest extending fiver hertz from each side of the peak. If there is a need to focus on a region, click on the highlight icon to deactivate it.Then left click and drag to zoom in on the region. Click on the highlight icon again to make the integration function active and move to the next peak. Left click and drag the cursor through the integral.Click the Return Save Regions button. Shown here is a representative spectrum of the Carbon NMR analysis in the carbonyl carbon region. The NMR signals of EPA and DHA on the sn-1, 3 and sn-2 positions are shown.These signals can be used for the quantitative determination of EPA and DHA. Shown here is a representative spectrum of the proton NMR analysis. The NMR signals of EPA and DHA that can be used for their determination are shown.The proton NMR spectrum is characterized by a narrower spectral width as compared to the carbon NMR spectrum and thus a lower spectral resolution. Once mastered, this technique can be done in less than 30 minutes if it is performed properly. After its development, this technique paved the way for researchers in the field of lipid analysis to rapidly analyze various lipid components that appear in multi-component matrices, such as dietary supplements, vegetable oil, and biofuels.After watching this video, you should have a good understanding of how to apply proton and carbon, NMR spectroscopy for the analysis of the lipid profile of fish oil supplements. Don't forget that working with strong magnetic fields produced by NMR spectrometers can be extremely hazardous to pacemakers and surgical protheses as well as electronic items such as credit cards and watches.

Research

JoVE Journal

Chemistry

Isolation and Chemical Characterization of Lipid A from Gram-negative Bacteria

Aislamiento y Caracterización química del lípido A de bacterias gram-negativas

Lipopolysaccharide (LPS) is the major cell surface molecule of gram-negative bacteria, deposited on the outer leaflet of the outer membrane bilayer. LPS ...

Investigación

Química

Structure and Coordination Determination of Peptide-metal Complexes Using 1D and 2D 1H NMR

Structure and Coordination Determination of Peptide-metal Complexes Using 1D and 2D 1H NMR

Copper (I) binding by metallochaperone transport proteins prevents copper oxidation and release of the toxic ions that may participate in harmful redox ...

Methods to Identify the NMR Resonances of the 13C-Dimethyl N-terminal Amine on Reductively Methylated Proteins

Methods to Identify the NMR Resonances of the 13C-Dimethyl N-terminal Amine on Reductively Methylated Proteins

Nuclear magnetic resonance (NMR) spectroscopy is a proven technique for protein structure and dynamic studies. To study proteins with NMR, stable magnetic ...

HKUST-1 as a Heterogeneous Catalyst for the Synthesis of Vanillin

HKUST-1 como un catalizador heterogéneo para la Síntesis de la vainillina

Vanillin (4-hydoxy-3-methoxybenzaldehyde) is the main component of the extract of vanilla bean. The natural vanilla scent is a mixture of approximately ...

Spin Saturation Transfer Difference NMR (SSTD NMR): A New Tool to Obtain Kinetic Parameters of Chemical Exchange Processes

Spin Saturation Transfer Difference NMR SSTD NMR: A New Tool to Obtain Kinetic Parameters of Chemical Exchange Processes

Transferencia de giro Saturación Diferencia RMN (MSITS RMN): Una nueva herramienta para obtener los parámetros cinéticos de los procesos de intercambio químico

This detailed protocol describes the new Spin Saturation Transfer Difference Nuclear Magnetic Resonance protocol (SSTD NMR), recently developed in our ...

Insights into the Interactions of Amino Acids and Peptides with Inorganic Materials Using Single-Molecule Force Spectroscopy

Miradas en torno a las interacciones de los aminoácidos y péptidos con materiales inorgánicos Uso de una sola molécula de la Fuerza Espectroscopia

The interactions between proteins or peptides and inorganic materials lead to several interesting processes. For example, combining proteins with minerals ...

Coulomb Explosion Imaging as a Tool to Distinguish Between Stereoisomers

Imágenes de explosión de Coulomb como herramienta para distinguir entre los estereoisómeros

This article shows how the COLTRIMS (Cold Target Recoil Ion Momentum Spectroscopy) or the "reaction microscope" technique can be used to distinguish ...

Electrochemical Impedance Spectroscopy as a Tool for Electrochemical Rate Constant Estimation

Espectroscopia de impedancia electroquímica como herramienta para la valoración constante de tipo electroquímico

Electrochemical impedance spectroscopy (EIS) was used for advanced characterization of organic electroactive compounds along with cyclic voltammetry (CV). ...

Color Spot Test As a Presumptive Tool for the Rapid Detection of Synthetic Cathinones

Prueba color como una herramienta presuntiva para la detección rápida de sintético Cathinones

Synthetic cathinones are a large class of new psychoactive substances (NPS) that are increasingly prevalent in drug seizures made by law enforcement and ...

A New Straightforward Method for Lipophilicity (logP) Measurement using 19F NMR Spectroscopy

A New Straightforward Method for Lipophilicity logP Measurement using 19F NMR Spectroscopy

Un nuevo método sencillo para la medida de lipofilicidad (logP) 19F NMR espectroscopia

Fluorination has become an effective tool to optimize physicochemical properties of bioactive compounds. One of the applications of fluorine introduction ...