TOF-SIMS imaging of chlorhexidine-digluconate transport in frozen hydrated biofilms of the fungus Candida albicans

The diffusion of the anti-microbial chlorhexidine digluconate (CHG) has been studied in C. albicans biofilms by time-of-flight secondary-ion mass spectrometry (TOF-SIMS). C. albicans has been shown to become resistant to common anti-microbial agents, including CHG, when growing as a biofilm. Mass transport resistance within biofilms has commonly been suggested as a resistance mechanism, but measurement of transport for most anti-microbial agents in biofilms has proven extremely difficult because of the heterogeneity of the biofilms and the difficulty in detecting these agents within an intact biofilm. In this study, TOF-SIMS has been used to study the transport of CHG and glucose in a frozen hydrated biofilm. The TOF-SIMS images reveal a progression of CHG from the top of the biofilm to its base with time. Images suggest that there are channels within the biofilm and show preferential binding of CHG to cellular components of the biofilm. Additionally, both living and dead cells can be identified in the TOF-SIMS images by the sequestration of K⁺ and the presence of cell markers. This study demonstrates that TOF-SIMS has the unique potential to simultaneously observe the presence of an antimicrobial agent, concentration of nutrients, and the viability of the cell population.     

Rapid biomass analysis

New, rapid, and inexpensive methods that monitor the chemical composition of corn stover and corn stover-derived samples are a key element to enabling the commercialization of processes that convert stover to fuels and chemicals. These new techniques combine near infrared (NIR) spectroscopy and projection to latent structures (PLS) multivariate analysis to allow the compositional analysis of hundreds of samples in 1 d at a cost of about $10 each. The new NIR/PLS rapid analysis methods can also be used to support a variety of research projects that would have been too costly to pursue by traditional methods.     

Templateless assembly of molecularly aligned conductive polymer nanowires: a new approach for oriented nanostructures

Although oriented carbon nanotubes, oriented nanowires of metals, semiconductors and oxides have attracted wide attention, there have been few reports on oriented polymer nanostructures such as nanowires. In this paper we report the assembly of large arrays of oriented nanowires containing molecularly aligned conducting polymers (polyaniline) without using a porous membrane template to support the polymer. The uniform oriented nanowires were prepared through controlled nucleation and growth during a stepwise electrochemical deposition process in which a large number of nuclei were first deposited on the substrate using a large current density. After the initial nucleation, the current density was reduced stepwise in order to grow the oriented nanowires from the nucleation sites created in the first step. The usefulness of these new polymer structures is demonstrated with a chemical sensor device for H(2)O(2), the detection of which is widely investigated for biosensors. Finally, we demonstrated that controlled nucleation and growth is a general approach and has potential for growing oriented nanostructures of other materials.     

Average peptide score: a useful parameter for identification of proteins derived from database searches of liquid chromatography/tandem mass spectrometry data

The quantity and variable quality of data that can be generated from liquid chromatography (LC)/mass spectrometry (MS)-based proteomics analyses creates many challenges in interpreting the spectra in terms of the actual proteins in a complex sample. In spite of improvements in algorithms that match putative peptide sequences to MS/MS spectra, the assembly of these lists of possible or probable peptides into a 'correct' set of proteins is still problematic. We have observed a trend in a simple relationship, derived from standard database search outputs, which can be useful in assessing the quality of a MS/MS-based protein identification. Specifically, the ratio of the protein score and number of non-redundant peptides, or average peptide score (APS), can facilitate initial filtering of database search results in addition to providing a useful measure of confidence for the proteins identified. This parameter has been applied to results from the analysis of multi-protein complexes derived from pull-down experiments analyzed using a two-dimensional LC/MS/MS workflow. In particular, the complex list of protein identifications derived from a drug affinity pull-down with immobilized ampicillin and an E. coli lysate was greatly simplified by applying the APS as a filter, allowing for facile identification of the penicillin-binding proteins known to interact with ampicillin. Furthermore, an APS threshold can be used for any data sets derived from electrospray ionization (ESI)- or matrix-assisted laser desorption/ionization (MALDI)-MS/MS experiments and is also not specific to any database search program.     

Octylgermane on gold: synthesis, oxidation, and pattern formation

Chemisorbed monolayers of octylgermane (C8H17GeH3) on gold have been formed by vapor deposition in ultrahigh vacuum. The monolayer has been characterized by X-ray photoelectron and reflection absorption infrared spectroscopies (XPS and RAIRS) and scanning tunneling microscopy (STM). XPS data indicate the monolayer can be oxidized by exposure to ozone. STM images exhibit a complex pattern which can be modeled as strain-mediated spinodal decomposition.     

Systems-based design of bi-ligand inhibitors of oxidoreductases: filling the chemical proteomic toolbox

Genomics-driven growth in the number of enzymes of unknown function has created a need for better strategies to characterize them. Since enzyme inhibitors have traditionally served this purpose, we present here an efficient systems-based inhibitor design strategy, enabled by bioinformatic and NMR structural developments. First, we parse the oxidoreductase gene family into structural subfamilies termed pharmacofamilies, which share pharmacophore features in their cofactor binding sites. Then we identify a ligand for this site and use NMR-based binding site mapping (NMR SOLVE) to determine where to extend a combinatorial library, such that diversity elements are directed into the adjacent substrate site. The cofactor mimic is reused in the library in a manner that parallels the reuse of cofactor domains in the oxidoreductase gene family. A library designed in this manner yielded specific inhibitors for multiple oxidoreductases.     

Determination of capsaicinoids in salsa by liquid chromatography and enzyme immunoassay

Two simple and rapid methods were developed to monitor pungency of salsa in production. Capsaicin (C) and dihydrocapsaicin (DHC) were quantitated in 17 commercially available tomato-based salsas by enzyme immunoassay (EIA) and liquid chromatography (LC) with fluorescent detection. Samples were extracted with methanol and the extracts were subjected to solid-phase extraction (SPE) using polystyrene-divinylbenzene columns. Analysis of SPE eluates showed good correlation (r2 = 0.953) between LC and EIA, with a slightly high bias for EIA. Salsa fortified with C and DHC from 0.118 to 103.2 microg/g resulted in recoveries of 90-112% (C) and 76-97% (DHC). Limits of detection by LC were 0.1 microg/g for each capsaicinoid and 0.1 microg/g by EIA for total capsaicinoids. The LC on-column response was linear from 0.2 to 100 ng for both C and DHC, whereas the working range for EIA was 0.1-2.0 ppm. Pungency varied between different salsa brands labeled mild, medium, and hot.     

POSTIRRADIATION PROPERTIES OF A UV-SENSITIVE VARIANT OF CHO

Abstract A UV-hypersensitive mutant of Chinese hamster ovary (CHO) cells, termed 43-3B, has been used in a comparative study with the wild type CHO in order to determine the involvement of repair in several postirradiation phenomena. 43-3B has the same growth rate and chromosome number as the wild type CHO-9. It is hypersensitive to UV irradiation (D₀ of 0.3 J/m² as compared to 3.2 J/m² for the wild type). 43-3B shows only about 17% of the UV-stimulated unscheduled DNA repair synthesis of CHO-9 as measured by autoradiography. When breaks in supercoiled chromatin are measured after UV by the nucleoid sedimentation method, the mutant appears to be capable of carrying out only limited incision. A much reduced ability to recover control rates of semiconservative DNA synthesis after UV irradiation was observed in the repair-deficient 43-3B cell line, suggesting that the removal of UV-induced replication blocks by excision repair is the most important factor in allowing recovery of UV-inhibited DNA synthesis. Recovery of colony-forming ability between fractionated UV exposures was observed in the wild type CHO-9, but little recovery was seen in 43-3B. This indicates that excision repair capability can also be important in split-fluence recovery.     

Cholesteric liquid crystal displays as optical sensors of barbiturate binding

The influence on the optical properties of cholesteric liquid crystal displays (LCDs) was examined for neutral molecule binding by mesogen/receptors in the mesomorphic phase. The motivation was to prepare neutral molecule sensors that use a colour change to signal analyte binding. A receptor that binds barbiturate analytes was modified with two or one cholesteryl groups to yield compounds 2 and 3, respectively. LCDs were prepared by incorporating one of the receptor/mesogen compounds into a cholesteric LC blend along with a potential H‐bonding guest. The optical properties of the LCDs were then determined by measuring the absorbance of the displays. For various LCDs, the colour of the display depended upon several factors: the amount of guest molecule used, the number of cholesteryl side chains on the receptor and the mole concentration of receptor/mesogen in the blend. In particular, complementary host/guest binding of H‐bonding analytes by the bis(cholesteryl) receptor 2 in a cholesteric LCD caused a change of up to +70 nm, which was observed by the naked eye as a blue‐to‐orange colour change. Control experiments confirm that the colour of an LCD is a consequence of molecular recognition in the mesomorphic phase.