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Minute Papers Due 10/05/2012

Please post this week's minute papers as "comments" to this post. Minute papers should be posted by 5 pm on Friday. Feel free to read your classmate's posts.


Minute Paper #5 (10/03/2012) – Sarah Anciaux

Title: Localization and Quantization of Drugs in Animal Tissue by use of Desorption Electrospray Ionization

Author: Zhiyang Zhoa et al.

Jounral: Analytical Chemistry

In this paper the authors attempt to develop a new method for determining the location and amount of drugs present in animal brain tissue, under ambient conditions. The use of desorption electrospray ionization (DESI) allows for ambient conditions and minimal sample preparation.

The authors developed a quantitative ambient detection method for drugs in animal tissue by implementation of a DESI source coupled to an orbitrap high resolution mass analyzer. The DESI source allows for easier 2D imaging due to the ambient conditions and solid samples. By placing a rat brain cross section on the DESI stage they are able to optimize the source for the drug placed on the rat brain. After optimization the authors dosed mice with the drug and quantified the amount of drug seen in the brain with a 2D scan. This was done by mapping the specific m/z ratios of interest. By making calibration curves and using an internal standard they were able to turn a usually only semi-quantitative method into a quantitative one. The authors also ensured the final quantization of drug in the rat brain by checking the concentration determined by the calibration curve against the concentration found upon drug extraction and LC-MS-MS analysis.

The authors claim to have successfully developed and implemented an ambient drug imaging and detection platform in animal tissue, but there are some areas that I think could be addressed further. The authors claim that they have developed a drug detection platform but by only testing one type of drug I question the validity of the statement. By only investigating one drug it is possible that this platform will not work well for many other types of drugs. The desorption mechanism of DESI generally requires that the sample be dissolved in the solvent spray in order to desorb, so if a drug of interest either somehow binds more in the tissue or is not easily dissolved then quantification of that drug would be very difficult as it would be hard to desorb. The authors also describe the use of an internal standard that is very similar to the drug of interest. This is done so that the ionizations are similar for better quantification. This would mean that for every drug of interest you would need to find a new internal standard of similar structure to the drug of interest. This would be very time consuming if trying to look at multiple drugs in a tissue at one time. It would be beneficial if a more universal standard could be utilized. The use of an internal standard also draws some criticism as it is possible the addition of the internal standard might somehow alter the sample or drug of interest so further investigation into the effects of this might be necessary.

Minute Paper #5–Marzieh Ramezani

Title: Thermal Processing as a Means to Prepare Durable, Submicron Thickness Ionomer Films for Study by Transmission Infrared Spectroscopy

Author: Carol Korzeniewski

Journal: Analytical Chemistry

Fuel cells have attracted a lot of attentions recently because of being efficient, clean and providing high electric power. The major challenge to develop them is improving properties of ion conductive medium which separate anode and cathode and it is composed of perfluorosulfonic acid ionomers and Nafion membranes. In this study, high temperature solution processing has been used to make highly durable, ultrathin, and freestanding fuel cell membranes which their structure properties have been studied by employing transmission infrared spectroscopy.

Vibrational modes of Nafion membrane which have been casted at 23 ºC and 150 ºC were collected. The differences were investigated, their bands were assigned and density function theory (DFT) was applied to examine molecular structures and confirm assignment of the experimental bands. For example; in both spectras the bands for C-F stretch (1155 cm-1), sulfonate and ether side chains were observed. By observing some differences of bands in two set of spectra such as band broadening (1070-1250 cm-1) and using computational predictions, it was concluded that the dispersed ionomer at high temperatures adopts more extended structure relative to the room temperature one and upon cooling to ambient temperature forms highly crystalline hydrophobic domains.

The author has claimed that using IR spectroscopy for investigation of nafion membranes is simple, non-destructive and can provide good insight about functional groups in its surrounding environment. I think using Raman spectroscopy would be helpful because it is also really non-destructive method. IR spectroscopy cannot be used to obtain the quantitative results. In the other hand, the structure of fuel cell membranes is complex and assigning IR spectra of complex compounds have a huge uncertainty. Also, a lot of compounds are not IR active. Simple techniques such as 1H NMR or 19F NMR might be good choices to study them too.

Title: Infrared Spectroscopy of the Mass 31 Cation: Protonated Formaldehyde vs Methoxy
By: M. A. Duncan et al.
Journal: The journal of physical chemistry A

In this paper, the group studied the structure of the mass 31 cation, [C,H3,O]+. The proposed cations were CH2OH+ protonated formaldehyde and CH3O+ methoxy. They could not find the evidence of existence of H2OCH+ oxonio-methylene, predicted from computational chemistry. They mentioned that studying structure of ions is important for mass spectroscopy. It would help understanding the plasma phase, which is crucial for interstellar space. For this study, they broadened ions in a supersonic beam, mass-selected in a time-of-flight spectrometer, and used infrared laser photodissociation spectroscopy. They also used computational methods using the Gaussian03 program package. They used the MP2/cc-pVTZ level of theory for energetics and the DFT/B3LYP for vibrational spectra.
They said there are three perspectives for mass 31 cation, which I mentioned above. Protonated formaldehyde has a singlet ground state and the most stable one among them. Methoxy is in a triplet ground state and the second stable one. The energy of methoxy is 96.0 kcal/mol higher than that of protonated formaldehyde. Thus, it is hard to get methoxy because of its high energy. They studied IR spectra of ions and investigated the method to get methoxy. In IR spectra, they reported the highest 3182cm-1 peak is for O-H stretch vibration, evidence of dominating protonated formaldehyde. They said 2469, 2717, and 2747 cm-1 peaks are for methoxy. They compared their results with computational works, and they fit well. They integrated the IR peaks to get relative amount of ions by comparing area. They studied the influence of precursors of the cations. For methanol precursor, the methoxy:protonated formaldehyde ratio was 1:53 and for ethanol precursor, the ratio was 1:13. They changed the discharge/expansion conditions when they produced ions. To get more methoxy, they needed to cool down the plasma quickly to prevent ions from crossing barriers. They compared 25K and 315K. At 25K, the methoxy:protonated formaldehyde ratio was 1:12 and , at 313K, the ratio was 1:34.
Their method was good to study the minor species and the structures of them. However, there are points that could be improved. First of all, they said they could not find any evidence for oxonio-methylene. If they use even lower temperature then 25K, they can freeze ionic species and detect oxonio-methylene. For quantitating method, integrating the area under the IR peaks, I am not sure about this method. In case of protonated formaldehyde, it corresponds to only one peak, so integrating would work. However, in case of methoxy, it has three corresponding peaks and summing those peaks is not comparable with protonated formaldehyde which has only one peak. Integrating the IR peaks is good for brief insight but not good for accurate value. They could investigate the way measuring NMR using plasma source. If ethanol gives the high ratio of methoxy, they could try higher alcohol, such as isopropanol and butanol. In addition, they can study other ion species, such as C6H5+. They might also be able to study anions using this technique.

Title: Consequences of Interfacial Viscoelasticity on Thin Film Stability
Authors: L. Rosenfeld et al.
Journal: Langmuir

In this publication, L. Rosenfeld et al. detail their work on the study of the surface viscoelasticity of three separate surfactants in thin films of water. The authors performed these studies to gain insight into how surface viscoelasticity affects the stability of tear films on the human eye. The three surfactants that were studied were arachidyl alcohol (AA), 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC), a slightly viscoelastic lipid, and meibum, a strongly viscoelastic lipid found in tear films. To understand these thin film systems, the authors employed three separate experimental techniques: interfacial shear rheometry (ISR), laser interferometry, and dewetting experiments.

In the ISR experiments, the surface elastic and viscous moduli of monolayers of aqueous solutions of each of the surfactants were measured at different surface pressures. The AA had a fairly constant elastic modulus and very low viscous modulus values, indicating that AA formed a Newtonian monolayer. For DPPC, the moduli values were below the sensitivity of the rheometer at surface pressures below 30 mN/m. At surface pressures above 30 mN/m, both moduli began to increase as a function of surface pressure, with the viscous surface modulus being orders of magnitude above the elastic modulus. The meibum showed significant viscoelasticity with both moduli growing over three orders of magnitude over a range of surface pressures from 0 to 40 mN/m. The elastic modulus dominated at higher surface pressures.

Laser interferometry was performed on droplets of the solutions to determine the droplet size and shape. Both the AA and DPPC droplets had a spherical shape, with the AA droplet having the largest thickness. The meibum droplet was fairly wrinkled and featured a crater in the center. The authors rationalized this morphology as the result of the high viscoelasticity of the meibum.

Dewetting, which is when the surfactant droplet changes from a spherical shape to a flat shape and the film breaks up, occurs when some of the solution evaporates and the droplet shrinks to a critical thickness. To observe this phenomenon, the authors filmed the dewetting process of each surfactant droplet at various surface pressures. At a surface pressure of 15 mN/m, the AA was unable to stabilize the film. However, both DPPC and meibum were able to inhibit the dewetting of the droplet and stabilized the droplet for over 30 seconds. At a surface pressure of 25 mN/m, the stabilization of each of the surfactants improved. In fact, the meibum completely stabilized the droplet for tens of minutes.

While the publication is very solid, there were several aspects of the experiments that are questionable. For instance, because the temperature of each of the experiments is never stated, it is assumed they were performed at room temperature. However, the experiments should be performed at 35 C, the temperature of the human eye, in order to truly simulate human tear films. Additionally, the authors fail to discuss how the thin film behavior of these surfactants differs from the bulk behavior. This sort of analysis would underscore the effect of surface tension at these small film thicknesses.

Sarah Gruba
Minute Paper 5
Article: Noninvasive Imaging of Intracellular Lipid Metabolism in Macrophages by Raman Microscopy in Combination with Stable Isotopic Labeling
Authors: Matthaus, C., Et.al
Journal: Analytical Chemistry

Macrophages play an important role in the innate immune system by phagocytosing extracellular junk and destroying pathogen infected cells. In high cholesterol diets these cells have the potential to not receive or detect a signal to stop uptaking extracellular fluids and will put the extra lipids and cholesterol in cytosolic organelles. The macrophage then takes on a foamy like appearance which is believed to be one of the first stages of atherosclerosis. This study set out to look at how these foam cells actually uptake lipids and cholesterol and spread them throughout the cell. The authors relied on Raman microscopy along with deuterium labeling and kits/assays that measured the amount of proteins, triglycerides, or cholesterol using absorption.

Initially, monocytes were cultured until mature macrophages were formed. Some of these macrophages were separated and incubated with palmitic acid or oleic acid. After washing, a portion of the cells went to a BCA Assay to quantify the total protein or a triglyceride kit to determine total triglyceride in the cell. Another portion of the cells were incubated with triglycerides and then put in the triglyceride kit, and the rest were fixed and grown on calcium chloride slides for Raman scattering.

The other mature macrophages were incubated with cholesterol for 1, 2, or 3 days. A portion of them were put in a cholesterol content kit per instructions and a BCA Assay to normalize the cholesterol with the amount of proteins in each sample. The rest of the sample was fixed and grown on calcium chloride slides for Raman scattering.

Using the Raman intensities in each spot of the cell they could integrate the data, to create an image of the cell showing were all the deuterated molecules they incubated with (cholesterol, palmitic acid, or oleic acid) were stored in the cell. The cells were found to uptake fatty acids at a much greater rate than the cholesterol, depositing them both in lipid droplets. However, unlike the fatty acids, the cholesterol did not increase in the cell after 24 hours of incubation. It also consistently showed that the cholesterol was being molecularly changed, seen by the spectral shift, which did not happen to the fatty acids. I think another interesting component that they could look at is how the change in the cell affects the release of cytokines. This would be interesting because if it increases the release, it would mean more macrophages and other immune cells would be called to the area causing vascular restriction and possibly platelet clotting. Or they could potentially use a microfluidic devise to see if the cells will actively go up the cholesterol gradient and uptake more cholesterol, or change the amount of cholesterol in their own experiment to see if macrophages can only uptake a certain amount or can increase the amount it takes in.

Minute Paper #5 October 3, 2012

Title: Ultrasensitive Liquid Chromatography-Tandem Mass Spectrometric Methodologies for Quantification of Five HIV-1 Integrase Inhibitors in Plasma for a Microdose Clinical Trial

By: Li Sun, Hankun Li and Kenneth Willson

Integrases are a class of enzymes that insert genome sequences into a host cell’s DNA. Integrase inhibitors can act as treatments for viral diseases by blocking the activity of this enzyme, which stops the spread of the virus since integration is a necessity for viral replication. Due to this, HIV-1 integrase strand transfer is used as a clinical treatment for HIV-1 infection. The chosen technique for drug candidate screening is microdosing, in which less than 1% of a pharmaceutical is administered, or no more than 100 μg. This technique has the benefit of being safe and presenting minimal adverse side effects due to the small dose, however it requires bioanalytical assays with a concentration range of pg/mL, an extremely sensitive capacity.

Accelerator mass spectrometry (AMS) is the current method used in assisting microdosing. Different from regular mass spectrometry, AMS accelerates ions to exceptionally high kinetic energies before performing mass analysis on each species. This technique has the advantage of being able to detect radio isotopes that are long lived and naturally occurring, however it requires specific radiolabeled compounds and can be used in only certain facilities. The authors in this paper seek to develop automated liquid chromatography – mass spectrometry/ mass spectrometry (LC-MS/MS). By attempting to quantify integrase inhibitors within plasma samples, the limits of detection can be observed and this technique can be evaluated as a suitable replacement for AMS.

The integrase inhibitor Raltegravir was the specific species of interest in this study due to its potency and currently being the only drug in this class approved for HIV-1 infection treatment. Double extractions were performed on all assays during sample preparation and analysis used ultra performance liquid chromatography (UPLC) coupled with MS/MS which had a negative ion electrospray introduction. Results indicated a 1 pg/mL limit of quantification for raltegravir and a 2 pg/mL limit for the other four proprietary compounds investigated. Using a standard curve concentration range, the accuracy was determined to be 93.8-107% and a precision of 1.2-14.1% was achieved.

The results of this study indicated that LC-MS/MS can be applied to microdosing in an accurate and reproducible manner. This technique has the advantage of being cost effective while eliminating the need for radiolabeled compounds over traditional AMS. Problems arose which are typical of chromatography experiments such as isobaric interferences, the matrix effect and high levels of chemical background interferences. Although the authors minimized the matrix effect by employing a negative ion electrospray introduction to MS/MS analysis, they did not perform any additional experiments to attempt to further minimize this effect. Altering the pH of the mobile phase when using UPLC has been shown to decrease the matrix effect^1, so I would run some experiments changing the mobile phase to attempt to decrease the matrix effect and possibly even diminish some chemical background interferences.

1. Chambers, E., Lu, Z., & Wagrowski-Diehl, D. M. (2007). Systematic and Comprehensive Stragey for Reducing Matix Effects in LC/MS/MS Analyses. ScienceDirect: Journal of Chromatography B 852, 22-34.

Title: Chiroptical Sensing of Citronellal: Systematic Development of a
Stereodynamic Probe Using the Concept of Isostericity

By: Wolf et.al.

Journal: Chem Comm

Link: http://pubs.rsc.org/en/content/articlepdf/2012/cc/c2cc36267h

In this paper, the authors designed a stereodynamic induced circular dichroism (ICD) probe for enantioselective sensing of citronellal. This is accomplished with a arylacetylene-based dialdehyde probe. (See the paper for the molecule structures) The idea, I think, is that upon interaction with the target molecule of interest, the probe molecule will react with the target and produce an axially chiral product.

The authors synthesized their molecule (bisaniline 2) themselves, but I don’t really want to get into that. (sorry organic chemists) What is interesting about their molecule is that it is prochiral in a sense. So when they just have it in solution, it is CD silent, meaning it does not bend light. The authors believe this is due to the fact that bisaniline 2 adopts rapidly interconverting chiral and achiral formations. So it then has no “chirality.” Then when they add citronellal, the axially chiral diimine product forms and then they got a CD readout, now that the chiral molecule was in solution. From this, they can quantitatively deduce the amount of enantiopure citronellal based on the intensity of the CD spectra peaks.

So it is a pretty cool way to detect enantiopurity, which is important especially in drug discovery and delivery. It is also nice since the reaction is clean and no further steps are needed before the CD spectra is taken. I do however wonder how well. This sensor reacts with other target molecules, like acetaminophen for example. Will it still work the same way? Is it a universal sensor, or do they have to design and synthesize another probe molecule each time? I suspect it isn’t universal, so it might not be as useful as I had hoped when I first started reading this paper.

Title: Design and Development of a Field Applicable Gold Nanosensor for the Detection of Luteinizing Hormone
Author: Ajit Zambre, et al.
Journal: Analytical Chemistry

In this paper, the authors developed a new strategy for sheep Luteinizing Hormone (LH) detection by using peptide conjugated gold nanoparticles (AuNP-LHP). Detection was achieved through the competitive binding to the sheep antibody of LH (anti-LH) between AuNP-LHP and LH in the analysis sample. The main techniques utilized in this paper involved physicochemical analysis of AuNP-LHP, including UV-visible absorption spectrum, TEM analysis and disc centrifuge sedimentation analysis, and Enzyme-linked immunosorbent assay (ELISA). For ELISA, the analyte is usually an antigen, which is attached to a solid surface. Specific antibody which is linked to an enzyme is then applied to the surface and binds to the antigen. Finally, the enzyme’s substrate is added to react with the enzyme and produces signals for detection.

As to the experiment, a peptide sequence with thiol functional group at the N-terminus of sheep LH (LPH) was synthesized and conjugated to AuNPs by the exchange reaction with thiol containing polyethylene glycol, which was first coupled to the AuNPs, since direct conjugation of AuNPs with LHP would cause aggregation of AuNPs. Subsequent physicochemical analysis of AuNP-LHP indicated the conjugation was highly stable. To test the specific binding of AuNP-LHP towards anti-LH, raised in rabbits, ELISA studies were performed and confirmed the binding affinities of AuNP-LHP towards anti-LH was similar to that of LH. This result also revealed that the biochemical function of LHP was not affected by the conjugation towards AuNPs. Thus AuNP-LHP could be used as a biosensor.

With these results, competitive immunostrip assay experiments were carried out to detect LH. 0.6 μg/μL (optimized concentration) of anti-LH was spotted on the nitrocellulose membrane and soaked in AuNP-LHP solutions with different concentration of LH (0, 10, 20, 50, 70, 100, 200, 300μg). The result showed that with no LH in the solution, red spot appeared on the membrane in less than 10 minutes caused by the binding of anti-LH and AuNP-LHP. However, no color change was observed when the concentration of LH was above 70 μg due to the stronger interaction between anti-LH and LH than that of anti-LH and AuNP-LHP. Field application of LH detection was also tested by using sheep’s blood, which was spiked with LH. Similar results were obtained with the detection limit of 46.66 ppm towards LH and further confirmed that AuNP-LHP could act as an effective biosensor.

In spite of the remarkable detection behavior towards LH shown by AuNP-LHP, this work can still be improved. As a field applicable sensor, it is almost impossible for the farmers to use newly synthesized AuNP-LHP for LH detection. Although AuNP-LHP can remain stable in biologically relevant solutions even after 24 hours, it is still hard to tell whether its binding affinity towards anti-LH would change during such a long time. So it is necessary to test the detection behavior of the AuNP-LHP again after keeping it for 24 hours. Also, to further validate the field application of AuNP-LHP, sheep’s blood containing LH instead of that is spiked with LH should be tested.

Title: Analysis of Lipids: Metal Oxide Laser Ionization Mass Spectrometry

Author: Casey R. McAlpin

Journal: Analytical chemistry

Casey R. McAlpin et al. demonstrates a new technique for rapid screening lipid detection using matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS). The application of MALDI MS for analysis of small molecules such as lipids has been limited due to interference from matrix peaks. Matrix-free systems have been explored to minimize this limitation. The surface properties of MgO and NiO are studied for matrix-free ionization of FAMEs, acylglycerides, glycolipids, and phospholipids. The results of other useful oxides as well as a control are reported.

Slurries of metal oxide powders were prepared for mass spectrometric analysis. Several metal oxides were evaluated for their ability to produce positive ions for methyl palmitate upon laser irradiation (NiO, MgO, FexOy, and ZnO). Among them NiO and MgO were the focus of lipid profiling because of their high signal-to-noise ratios. The author states that the contribution of photoelectric phenomena to ionization may not be the only factor influencing ionization activity for these metal oxides, and likely there is a combination of processes, although he does not explore this much. Previous studies have shown that activity is proportional to the surface area, and in some cases surface defects and increased surface area play a significant role by increasing the population of surface silanol groups which act as a proton source during ionization.

In the results the author shows that both MgO and NiO with a variety of particle sizes and from differing preparations had the same activity for MOLI MS, and thus deduces that surface area and surface defects do not play a significant role in ionization activity. Additionally, NiO and MgO with the highly defective primary surface did not produce ions, further supporting that surface defects are not critical to the ionization process.

For microbial lipid profiling, MOLI MS was applied to complex mixtures of lipids from vegetable oil shortening and bacterial and algal extracts, which were spotted directly onto NiO spots. The lipids consisted of individual colonies of E. coli and Algal. The peaks resulting from the lipids can be seen in the spectrum provided, and shows clear distinction between levels of unsaturation in comparison to other non-matrix methods. The author does acknowledge that quantitation using MOLI MS is difficult and further work needs to be done.

The author claims that surface do not affect ionization in these experiments, while other literature demonstrates they have. It would be interesting to look at the influence of defects inside the particles themselves (i.e. excessive oxygen or metal vacancies). The author also doesn’t discuss how the crystallinity of the metal oxide particles influences ionization. Crystallinity of metal oxides is known to have preferential electron affinity along different crystal axis and whether ionization of the particles transfers energy to the substrate or the species of interest could be dependant upon these properties.

Title: Ultra-Small, Highly Stable, and Sensitive Dual Nanosensors for
Imaging Intracellular Oxygen and pH in Cytosol

By: Wang et al.

Journal: Journal of the American Chemical Society

Although thin-film optical chemical sensors have proven to be viable analytical tools for chemical environment measurements, their planar bulky nature restricts them from in vivo use. Herein, small diameter nanoparticles functionalized with oxygen and pH sensitive probes were used as intracellular optical chemical sensors within rat kidney cells. The methods included characterization of synthesized materials, as well as fluorescence spectroscopy and confocal laser scanning microscopy.

Pluronic F-127 triblock copolymers, which are composed of a central hydrophobic chain flanked by two poly(ethylene glycol) hydrophilic chains with terminal FITC pH sensitive fluorophores, were mixed with hydrophobic luminescent oxygen probes (PtTPTBP) in acidic aqueous media. In addition, an inert hydrophobic reference probe and non-FITC labeled F-127 polymers were mixed in the media in order to prevent self-quenching of closely packed probes on both the interior and exterior of the nanoparticles. Silica condensation via TEOS addition was then completed about the micelles, which resulted in small particles (12 nm diameters) with internally labeled PtTPTBP oxygen probes and externally labeled FITC pH probes.

PtTPTBP’s sensitivity was demonstrated via fluorescence spectroscopy over a range of dissolved oxygen concentrations (0 - 41.25 mg/mL). Furthermore, Stern-Volmer plots illustrated a clear correlation between relative fluorescence and dissolved oxygen concentration. Normalized fluorescence intensity measurements suggested FITC emission properties are very sensitive to pH change, as FITC was determined to have pKa 6.4. This is indicative of the probe’s weakly acidic nature as well as the probe’s susceptibility to fluorescence intensity increase with deprotonation.

Although the authors provided novel materials methodology, I found their nanoparticles to have very little promise as widely used in vivo sensors. The nanoparticles could only enter rat kidney cells via electroporation, which thereby complicates in vivo sensing, but also negates any promise for the nanoparticles serving as diagnostic tools within living patients. Furthermore, many current silica materials are easily transported across cell membranes without electroporation, and this therefore makes the nanoparticles featured in this work less suitable for cellular work. Also, the authors mentioned signal intensity issues with NIR measurements of the oxygen probes. Extremely high concentrations of nanoparticles were required to obtain reasonable signal intensities, which reduced the effectiveness of cellular uptake. I also found little evidence that suggested respective locations of inert and active oxygen probes within the nanoparticle. This is a significant issue, because it is possible their assessment of dissolved oxygen within cells may be skewed by oxygen access discrepancies between the exterior and interior of the nanoparticle. Lastly, no complete stability studies of the probes or nanoparticles were conducted.

I think mesoporous silica nanoparticles (MSNs) would remedy some of the issues in this paper, as MSNs modified with oxygen sensitive probes would permit exterior and interior oxygen access. Furthermore, MSNs are easily taken up by cells without electroporation. Lastly, MSNs have long-term stability in biological media.

Title: Microheterogeneity of Some Imidazolium Ionic Liquids As Revealed by Fluorescence Correlation Spectroscopy and Lifetime Studies
Authors: Patra, S. and Samanta, A.
Journal: J. Phys. Chem. B

In this paper, the authors use fluorescence correlation spectroscopy to study the diffusion of probe molecules in certain imidazolium ionic liquids, concluding that there exist two distinct environments within the ionic liquid.

Ionic liquids have been of interest in recent years due to their properties, including low vapor pressure, high thermal stability and conductivity, nonflammability and the fact that other properties can be ‘tuned’ by selecting appropriate constituent ions. Previous studies concluded that ionic liquids are structured, and not homogeneous at the microscopic level.

The authors studied the diffusion through 1-alkyl-3-methyl-imidazolium based ionic liquids of three probe molecules: one cation, R123, and two neutral probes, DCM and 4NBD. The technique for this was fluorescence correlation spectroscopy, which measures the fluctuations of fluorescence intensity of a highly dilute solution to generate a correlation function. The decay of this correlation function gives information about dynamic molecular processes, including kinetic diffusion.

The decay of the correlation curves in the ionic liquids was fitted to a two-component diffusion model, showing that the probes exhibit a bimodal diffusion behavior, a fast diffusion mode and a slow diffusion mode. Because these probes exhibit single-component diffusion in conventional solvents, it was concluded that the bimodal diffusion behavior was a result of two distinct environments in the ionic liquids. The authors assigned these different diffusions to two different regions: domains formed by the hydrophobic alkyl tails on the imidazolium and domains around the ionic constituents of the ionic liquids. The authors also concluded that these two types of domain must be interconnected, ie, the hydrophobic alkyl region of one ionic liquid molecule must be connected with the hydrophobic alkyl region of another, and the same for the hydrophilic ionic regions. This continuity in the domain structure allows molecules to diffuse though only one type of domain, explaining the bimodal diffusion behavior.

Next, the authors studied the decay of the fluorescence of each diffusion mode. For every probe, the fluorescence decay was found to be biexponential, another result attributed to the heterogeneity of the ionic liquids. The neutral probes, DCM and 4NBD, are both known to have fast, short lifetime fluorescence decay in polar regions, so the authors attributed the fast DCM and 4NBD decays to molecules in polar regions, and slow decay to molecules in non-polar regions. By analyzing the weighted contribution of each component to the overall decay, the authors found that both of these molecules are distributed more in the long decay (non-polar) regions of the ionic liquid. The cationic probe, R123, is known to exhibit long-lifetime decay in polar regions and fast decay in non-polar regions. The weight of the long-lifetime decay was higher, so the authors concluded that R123 was mainly located in the polar environment. The authors used this result to explain the overall slower diffusion rate of R123 compared to the other probes, postulating that R123 experiences a strong dragging force due to hydrogen bonding and electrostatic interaction with the ionic constituents of the ionic liquids.

Title: Application of Operando XAS, XRD, and Raman Spectroscopy for Phase Speciation in Water Gas Shift Reaction Catalysts

By: Patolla et al.

Journal: ACS Catalysis

Catalysis often requires an understanding of the transient behavior of the catalyst structure and oxidation state as well as the identity and concentration of surface species during reaction. In light of this, the authors developed a system capable of performing operando X-ray adsorption spectroscopy (XAS), X-ray diffraction (XRD), and Raman spectroscopy to study the change of these properties during reaction. The authors chose to test this system by analyzing the water gas shift reaction, which produces CO2 and H2 from CO and H2O over pure and Cr2O3 doped gamma-Fe2O3 catalysts. gamma-Fe2O3 is hypothesized to reduce to catalytically active Fe3O4 during the water gas shift reaction1. To avoid deactivation of the catalyst due to sintering that can occur during this reduction, Cr2O3 is often used as a structural stabilizer for this catalyst1. The authors of this work aimed to use their system to better understand the role of the Cr2O3 dopant during the water gas shift reaction.
The X-ray absorption fine structure (EXAFS) spectra were measured for Fe2O3 on both the pure and the doped sample at ambient temperature under both O2 atmosphere and water gas shift reaction conditions. For both catalyst samples, the intensity of the peak associated with Fe-O interactions (at approximately 1.5 Å) decreased upon reaction; designating a reduction in the oxidation state of Fe characteristic of the formation of the catalytically active Fe3O4. Additionally, the intensity of the peak associated with Fe-Fe interactions (at approximately 3.0 Å) was found to decrease to a much greater extent for the Cr2O3 doped sample upon reaction; the authors to concluded that this difference would be explained if the doped sample is much more heterogeneous and disordered during reaction than the pure sample and, thus, would have a distribution of Fe-Fe bond distances. The Raman spectra collected at the same conditions indicated that alpha-Fe2O3 (signified by bands at 226 and 292 cm-1) formed under reaction in the pure samples, while no peaks corresponding to this transition were found in the spectra for the doped sample. This observation was also supported by the XRD diffraction patterns in which the alpha-Fe2O3 lines were much larger for the pure catalyst than for the doped catalyst. Based upon these observations, the authors concluded Cr2O3 inhibits the formation of alpha-Fe2O3 during reaction at room temperature which may be the cause for the reduced activity of pure gamma-Fe2O3.
The authors were successful in identifying the reduced formation of alpha-Fe2O3 in the doped catalyst during reaction at room temperature. The XRD patterns measured at 400 °C, however, demonstrated alpha-Fe2O3 is formed in the doped sample even in an O2 atmosphere prior to reaction. Raman measurements were not taken at this temperature due to radiation from the system’s heating unit. If this radiation could be contained, operando Raman measurements would provide a valuable tool in understanding what happens to the Cr2O3 on the catalyst surface at elevated temperatures, and may explain why doping is unable to inhibit the formation of alpha-Fe2O3 at these temperatures.

[1] Gonzalez, J. C.; Gonzalez, M. G.; Laborde, M. A.; Moreno, N. Appl. Catal. 1986, 20, 3−13.

Minute Paper #4
Tian Qiu 4651092

Title: Single Quantum Dot Based Nanosensor for Renin Assay
Journal: Analytical Chemistry
Authors: Yi Long, Chun-yang Zhang, et al.

In this article, the authors developed a single quantum dot (QD) based nanosensor for renin assay. Usually quantum dots (QDs) are semiconductors whose excitons are confined in all three spatial dimensions. Their features include size-dependent emission spectra, relatively high quantum yield and good resistance to chemicals, making it a great alternative to the organic fluorophores in fluorescence assay. Here the authors use QDs as the donor in fluorescence resonance energy transfer (FRET) and Cy5 as the acceptor.

The authors designed a QD/substrate/Cy5 complex. The substrate is a peptide sequenced as biotin-Lys-His-Pro-Phe-His-Leu-Val-Ile-His-Lys, and Cy5 is attached to the terminal lysine. They coated the QDs with streptavidin and then linked the biotin-linked substrate to the QD. Without the presence of renin, the fluorescence of Cy5 was observed because of the FRET between QDs and Cy5. After the addition of renin, only the fluorescence of QDs was observed because of the cleavage of the substrate by the renin proteolytic reaction.

Influence of Substrate-to-QD ratio on FRET efficiency was determined by both the single-QD-based nanosensor and the bulk measurement. In the single-QD-based measurement, a good linear correlation is obtained between the Cy5 counts and the Cy5-labeled substrato-to-QD ratio in the range from 1/1 to 48/1. The bulk measurement showed a similar result between the ratio ranges. Also they performed measurement of time profile of renin activity, indicating that the single-QD-based nanosensor is specific to renin. At last, the renin proteolytic assay was performed and the Michaelis-Menten expression is determined. The results showed that the kinetic parameters can be accurately evaluated by the single-QD-based nanosensor. The detection limit of this assay is as low as 25pM, which has improved by more than 40-fold compared with the method using EDANS/DABCYL as the fluorophore/quencher pair.

What I’m concerning about this method is whether this assay could be performed as real-time, in vivo or single-molecule. As indicated in the paper, QDs could be used for single-molecule assay, but this article didn’t show assays on single-molecule level. I would suggest use microarrays or microfluidics to perform single-molecule measurement. Also, if we could measure renin in vivo and real-time in the body, that would be exciting. But as QDs are usually composite of semiconductors like Cd and Se, which are poisonous to the body, they should find a way, like coating the QDs with biocompatible polymers while make sure the fluorescence efficiency is not affeted, to perform in vivo assays.

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