Oral squamous cell carcinoma diagnosed from saliva metabolic profiling.

Saliva is a noninvasive biofluid that can contain metabolite signatures of oral squamous cell carcinoma (OSCC). Conductive polymer spray ionization mass spectrometry (CPSI-MS) is employed to record a wide range of metabolite species within a few seconds, making this technique appealing as a point-of-care method for the early detection of OSCC. Saliva samples from 373 volunteers, 124 who are healthy, 124 who have premalignant lesions, and 125 who are OSCC patients, were collected for discovering and validating dysregulated metabolites and determining altered metabolic pathways. Metabolite markers were reconfirmed at the primary tissue level by desorption electrospray ionization MS imaging (DESI-MSI), demonstrating the reliability of diagnoses based on saliva metabolomics. With the aid of machine learning (ML), OSCC and premalignant lesions can be distinguished from the normal physical condition in real time with an accuracy of 86.7%, on a person by person basis. These results suggest that the combination of CPSI-MS and ML is a feasible tool for accurate, automated diagnosis of OSCC in clinical practice.

In situ monitoring of electrochemical reactions through CNT-assisted paper cell mass spectrometry.

A novel method of coupling electrochemistry (EC) with mass spectrometry (MS) is illustrated with a paper-based electrochemical cell supported by carbon nanotubes (CNTs). The electrochemically formed ions, created at appropriate electrochemical potentials, are ejected into the gas phase from the modified paper, without the application of additional potential. The electrochemical cell was fabricated by using a rectangular CNT-coated Whatman 42 filter paper with printed electrodes, using silver paste. This was used for studying the electrochemical conversion of thiols to disulfides, and the functionalization of polycyclic aromatic hydrocarbons (PAHs), which involve S-S and C-C bond formations, respectively. We also demonstrate the versatility of the set-up by utilizing it for the detection of radical cations of metallocenes, monitoring the oxidation of sulfides through the detection of reactive intermediates, and the detection of radical cations of PAHs, all of which occur at specific applied potentials. Finally, the applicability of this technique for qualitative and quantitative analyses of environmentally relevant molecules has been demonstrated by studying the electrochemical oxidation of glucose (Glu) to gluconic acid (GlcA) and saccharic acid (SacA).

Probing Coordination Complexes by Carbon Nanotube-Assisted Low-Voltage Paper Spray Ionization Mass Spectrometry.

Fragile transition metal complex ions such as [Cr(H2O)4Cl2]+, difficult to be observed by gas-phase spectroscopy, are detected easily with carbon nanotube (CNT)-assisted low-voltage ambient ionization mass spectrometry. Observation of various substituted ions with D2O and ROH (R = CH3, C2H5, ...) established the versatility of the technique in detecting diverse species. Ligand substitution occurring in solution was captured by the low-voltage technique. The extreme softness of the technique coupled with nanoscale ion sources enabled the creation of such species. Analysis was extended to other halides as well. The intensity of these fragile ions gradually disappeared at voltages beyond 500 V and are completely absent in standard high-voltage ionization. Detection of inorganic complexes further enhances the scope of low-voltage ionization.

Anisotropic Molecular Ionization at 1 V from Tellurium Nanowires (Te NWs).

Ionization of molecular species from one-dimensional (1D) tellurium nanowires (Te NWs) has been achieved at 1 V. Molecules with a range of chemical functional groups gave quality mass spectra with high signal/noise ratios and no fragment ions. Experiments suggest the possibility of emission of microdroplets of solution due to the intense fields at the ends or interfaces of nanostructures. It appears that electrolytic conduction of the solution wetting of the nanostructures and not the electronic conduction of the nanostructures themselves is involved in the ionization event. Anisotropy was seen when two-dimensionally aligned Te NWs were used for ionization. The orientation effect of aligned Te NWs on molecular ion intensity is demonstrated for many analytes including organic molecules and amino acids with experiments done using a silicon substrate having aligned Te NWs. These measurements suggest the possibility of creating a MS source that extends the applicability of mass spectrometry. Analysis of a variety of analytes, including amino acids, pesticides, and drugs, in pure form and in complex mixtures, is reported. These experiments suggest that 1D nanostructures in general could be excellent ionization sources.

Zero Volt Paper Spray Ionization and Its Mechanism.

The analytical performance and a suggested mechanism for zero volt paper spray using chromatography paper are presented. A spray is generated by the action of the pneumatic force of the mass spectrometer (MS) vacuum at the inlet. Positive and negative ion signals are observed, and comparisons are made with standard kV paper spray (PS) ionization and nanoelectrospray ionization (nESI). While the range of analytes to which zero volt PS is applicable is very similar to kV PS and nESI, differences in the mass spectra of mixtures are interpreted in terms of the more significant effects of analyte surface activity in the gentler zero volt experiment than in the other methods due to the significantly lower charge. The signal intensity of zero volt PS is also lower than in the other methods. A Monte Carlo simulation based on statistical fluctuation of positive and negative ions in solution has been implemented to explain the production of ions from initially uncharged droplets. Uncharged droplets first break up due to aerodynamics forces until they are in the 2-4 µm size range and then undergo Coulombic fission. A model involving statistical charge fluctuations in both phases predicts detection limits similar to those observed experimentally and explains the effects of binary mixture components on relative ionization efficiencies. The proposed mechanism may also play a role in ionization by other voltage-free methods.

Molecular ionization from carbon nanotube paper.

Ambient ionization is achieved by spraying from a carbon nanotube (CNT)-impregnated paper surface under the influence of small voltages (≥3 V). Organic molecules give simple high-quality mass spectra without fragmentation in the positive or negative ion modes. Conventional field ionization is ruled out, and it appears that field emission of microdroplets occurs. Microscopic examination of the CNT paper confirms that the nanoscale features at the paper surface are responsible for the high electric fields. Raman spectra imply substantial current flows in the nanotubes. The performance of this analytical method was demonstrated for a range of volatile and nonvolatile compounds and a variety of matrices.

Mutation-Dependent Refolding of Prion Protein Unveils Amyloidogenic-Related Structural Ramifications: Insights from Molecular Dynamics Simulations.

The main focus of prion structural biology studies is to understand the molecular basis of prion diseases caused by misfolding, and aggregation of the cellular prion protein PrPC remains elusive. Several genetic mutations are linked with human prion diseases and driven by the conformational conversion of PrPC to the toxic PrPSc. The main goal of this study is to gain a better insight into the molecular effect of disease-associated V210I mutation on this process by molecular dynamics simulations. This inherited mutation elicited copious structural changes in the ß1-α1-ß2 subdomain, including an unfolding of a helix α1 and the elongation of the ß-sheet. These unusual structural changes likely appeared to detach the ß1-α1-ß2 subdomain from the α2-α3 core, an early misfolding event necessary for the conformational conversion of PrPC to PrPSc. Ultimately, the unfolded α1 and its prior ß1-α1 loop further engaged with unrestrained conformational dynamics and were widely considered as amyloidogenic-inducing traits. Furthermore, the resulting folding intermediate possesses a highly unstable ß1-α1-ß2 subdomain, thereby enhancing the aggregation of misfolded PrPC through intermolecular interactions between frequently refolding regions. Briefly, these remarkable changes as seen in the mutant ß1-α1-ß2 subdomain are consistent with previous experimental results and thus provide a molecular basis of PrPC misfolding associated with the conformational conversion of PrPC to PrPSc.

A Spatial Model of Hepatic Calcium Signaling and Glucose Metabolism Under Autonomic Control Reveals Functional Consequences of Varying Liver Innervation Patterns Across Species.

Rapid breakdown of hepatic glycogen stores into glucose plays an important role during intense physical exercise to maintain systemic euglycemia. Hepatic glycogenolysis is governed by several different liver-intrinsic and systemic factors such as hepatic zonation, circulating catecholamines, hepatocellular calcium signaling, hepatic neuroanatomy, and the central nervous system (CNS). Of the factors regulating hepatic glycogenolysis, the extent of lobular innervation varies significantly between humans and rodents. While rodents display very few autonomic nerve terminals in the liver, nearly every hepatic layer in the human liver receives neural input. In the present study, we developed a multi-scale, multi-organ model of hepatic metabolism incorporating liver zonation, lobular scale calcium signaling, hepatic innervation, and direct and peripheral organ-mediated communication between the liver and the CNS. We evaluated the effect of each of these governing factors on the total hepatic glucose output and zonal glycogenolytic patterns within liver lobules during simulated physical exercise. Our simulations revealed that direct neuronal stimulation of the liver and an increase in circulating catecholamines increases hepatic glucose output mediated by mobilization of intracellular calcium stores and lobular scale calcium waves. Comparing simulated glycogenolysis between human-like and rodent-like hepatic innervation patterns (extensive vs. minimal) suggested that propagation of calcium transients across liver lobules acts as a compensatory mechanism to improve hepatic glucose output in sparsely innervated livers. Interestingly, our simulations suggested that catecholamine-driven glycogenolysis is reduced under portal hypertension. However, increased innervation coupled with strong intercellular communication can improve the total hepatic glucose output under portal hypertension. In summary, our modeling and simulation study reveals a complex interplay of intercellular and multi-organ interactions that can lead to differing calcium dynamics and spatial distributions of glycogenolysis at the lobular scale in the liver.

Teflon Spray Ionization Mass Spectrometry.

Polytetrafluoroethene, commonly known as Teflon, is a plastic famous for its inertness, strength, and nonstick properties, allowing its repeated use in many applications. We report the use of a triangularly cut Teflon substrate to take the place of paper in a form of spray mass spectrometry. A conducting wire (gold) at high potential (positive or negative) makes contact with a drop of the liquid sample at the apex of the triangle, causing a spray of droplets to be directed toward the heated inlet of a mass spectrometer. Saccharides, drugs, illegal additives, peptides, proteins, bilirubin, and vancomycin give mass spectra with high signal-to-noise (S/N) ratios, allowing detection at the nanogram per milliliter (ng/mL) level. Examination of each of these analytes demonstrates that Teflon spray is several orders of magnitude more sensitive than paper spray under the same conditions. Teflon spray ionization mass spectrometry is applied to the metabolomic and lipidomic profiling of biological fluid samples. Detection of polycyclic aromatic hydrocarbons is achieved with Teflon spray at 10 µg/mL concentrations. These experiments show the advantage of using Teflon over a normal paper substrate in detecting many environmentally and biologically relevant systems with high sensitivity and S/N ratio.

Studying reaction intermediates formed at graphenic surfaces.

We report in-situ production and detection of intermediates at graphenic surfaces, especially during alcohol oxidation. Alcohol oxidation to acid occurs on graphene oxide-coated paper surface, driven by an electrical potential, in a paper spray mass spectrometry experiment. As paper spray ionization is a fast process and the time scale matches with the reaction time scale, we were able to detect the intermediate, acetal. This is the first observation of acetal formed in surface oxidation. The process is not limited to alcohols and the reaction has been extended to aldehydes, amines, phosphenes, sugars, etc., where reaction products were detected instantaneously. By combining surface reactions with ambient ionization and mass spectrometry, we show that new insights into chemical reactions become feasible. We suggest that several other chemical transformations may be studied this way. This work opens up a new pathway for different industrially and energetically important reactions using different metal catalysts and modified substrate.