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Plant secondary compounds are diverse structurally, and associated biological effects can vary depending on multiple factors including chemical structure and reaction conditions. Phenolic compounds such as tannins can chelate dietary iron, and supplementation of animal species sensitive to iron overload with tannins may prevent/treat iron overload disorder. We assessed the nutrient and phenolic composition and iron-binding capacity of Carolina willow (Salix caroliniana), a plant fed to zoo-managed browsing herbivores. Based on studies in other plant species and the chemical structures of phenolic compounds, we hypothesized that the concentration of condensed tannins in willow would be inversely related to the concentration of phenolic glycosides and directly related to iron-binding capacity. Our results indicated that willow nutrient composition varied by year, season, and plant part, which could be taken into consideration when formulating animal diets. We also found that the predominant plant secondary compounds were condensed tannins with minimal phenolic glycosides. Instead of binding to iron, the willow leaf extracts reduced iron from the ferric to ferrous form, which may have prooxidative effects and increase the bioavailability of iron depending on animal species, gastrointestinal conditions, and whole animal processes. We recommend identifying alternative compounds that effectively chelate iron in vitro and conducting chelation therapy trials in vivo to assess potential effects on iron balance and overall animal health.

Iron overload disorder has been described in a number of zoo-managed species, and it has been recommended to increase the tannin composition of the diet as a safe way to minimize iron absorption in these iron-sensitive species. The goal of this study was to examine the potential of near infrared reflectance spectroscopy (NIRS) as a rapid and simple screening tool to assess willow (Salix caroliniana) nutrient composition (crude protein: CP; acid detergent fiber: ADF; neutral detergent fiber: NDF; lignin, gross energy: GE) and condensed tannin (CT) concentrations. Calibration equations were developed by regression of the lab values from 2 years using partial least squares on n = 144 NIRS spectra to predict n = 20 independent validation samples. Using the full 2-year dataset, good prediction statistics were obtained for CP, ADF, NDF, and GE in plant leaves and stems (r(2 ) > 0.75). NIRS did not predict lignin concentrations reliably (leaves r(2) = 0.52, stems r(2) = 0.33); however, CTs were predicted moderately well (leaves r(2) = 0.72, stems r(2) = 0.67). These data indicate that NIRS can be used to quantify several key nutrients in willow leaves and stems including concentrations of plant secondary compounds which, depending on the bioactivity of the compound, may be targeted to feed iron-sensitive browsing animals.

The fragmentation characteristics of five oligo-peptoids were studied under tandem mass spectrometry conditions. The charged peptoids were produced by protonation and alkali metal cation (Li + , Na + , K + , Rb + , and Cs + ) addition. The peptoids were ionized by the MALDI process and the resulting ions were fragmented via collision-induced dissociation (CID) experiments. All charged peptoids fragmented predominantly at the amide bonds. Highly abundant and sequence-dependent fragment ions were observed. The fragmentation patterns for the protonated peptoids and the metal cation adducts were strikingly different. All protonated peptoids fragmented by producing predominantly Y-type ions. The bias towards Y-ions was largely due to the greater proton affinity of the secondary amine at the terminal side of the Y-ions. All alkali metalated peptoids fragmented by producing both Y′- and B′-type ions, suggesting a “mobile metal cation” mechanism. For the peptoids with basic side chains, formation of the most abundant ions corresponded to the cleavage of the amide bonds at or near the basic residue. These results suggest that the metal cations are largely coordinated to the side chain of the basic residue. Chelation between the metal cation and the amino groups of the peptoids is an important factor to stabilize the fragment ions. For the peptoid without a basic side chain, the ion intensity was evenly distributed among all medium sized fragment ions. Fragmentations of protonated and alkali metalated peptoids yielded complementary sequential information, which demonstrated the practical utility of using mass spectrometry methods for de novo sequencing of peptoid libraries generated by combinatorial chemistry.

Differences in the fertilization behavior of Xenopus borealis from X. laevis and X. tropicalis suggest differences in the glycosylation of the egg jellies. To test this assumption, O-linked glycans were chemically released from the egg jelly coat glycoproteins of X. borealis. Over 50 major neutral glycans were observed, and no anionic glycans were detected from the released O-glycan pool. Preliminary structures of ∼30 neutral oligosaccharides were determined using matrix-assisted laser desorption/ionization (MALDI) infrared multiphoton dissociation tandem mass spectrometry (MS). The mass fingerprint of a group of peaks for the core-2 structure of O-glycans was conserved in the tandem mass spectra and was instrumental in rapid and efficient structure determination. Among the 29 O-glycans, 22 glycans contain the typical core-2 structure, 3 glycans have the core-1 structure and 2 glycans contained a previously unobserved core structure with hexose at the reducing end. There were seven pairs of structural isomers observed in the major O-linked oligosaccharides. To further elucidate the structures of a dozen O-linked glycans, specific and targeted exoglycosidase digestions were carried out and the products were monitored with MALDI-MS. Reported here are the elucidated structures of O-linked oligosaccharides from glycoproteins of X. borealis egg jelly coats. The structural differences in O-glycans from jelly coats of X. borealis and its close relatives may provide a better understanding of the structure-function relationships and the role of glycans in the fertilization process within Xenopodinae.

Anthrax infections progress at a rapid pace, making rapid detection methods of utmost importance. MALDI-MS proteomics methods focused on Bacillus anthracis detection have targeted chromosomally encoded proteins, which are highly conserved between closely related species, hindering species identification. Presented here is an AP-MALDI-MS method targeting plasmid-borne proteins from Bacillus spores for species-level identification. A bioinformatics analysis revealed that 60.3% and 75.4% of tryptic peptides from plasmid-borne proteins of B. anthracis and B. thuringiensis were species-specific, respectively. Reported here is a method in which plasmid-borne delta-endotoxins were extracted directly from B. thuringiensis spores in 100 mM KOH. The pH was then adjusted to 8 and a 5-min trypsin digestion was performed on the extracted proteins. The resulting tryptic peptides were analyzed by AP-MALDI-MS/MS, which produced a definitive identification the B. thuringiensis species-specific Cry1Ab protein with a MASCOT score of 278 and expect value of 7.5 x 10(-23). This method has demonstrated the detection and identification of B. thuringiensis spores at the species level following a 5-min trypsin digestion. The challenges in applying a similar approach to the detection of plasmid-borne protein toxins from B. anthracis are also discussed.

A nonenzymatic proteomics strategy is applied to the rapid identification of viruses. The approach provides solubilization and subsequent digestion of viral coat proteins in under 30 s. Acid digestions were carried out using a laboratory-quality microwave system equipped with temperature, pressure, and power controls, which allowed for precise optimization of experimental parameters. Under optimal conditions, this method provides an efficient alternative to traditional enzymatic digestion-based methods for virus identification. Following rapid microwave heating of a suspension of a model virus, RNA bacteriophage MS2, 13 chemical digestion products were detected in parallel with the coat protein precursor using MALDI-TOF MS. Because of the high sequence coverage obtained, the bacteriophage MS2 coat protein was identified with high confidence and the specificity of the identification allowed for the discrimination between bacteriophage MS2 and other closely related RNA bacteriophages.

A method for the rapid identification of Bacillus spores is proposed, based on the selective release and chemical digestion of small, acid-soluble spore proteins (SASPs). Microwave-assisted acid hydrolysis of SASPs from B. anthracis str. Sterne and B. subtilis str. 168 was accomplished in a single step requiring only 90 s of heating. The peptide products of the chemical digestion were identified by postsource decay sequencing with a MALDI-TOF-MS equipped with a curved-field reflectron. The specificity of the observed SASP peptides was evaluated using a cross-species sequence search. The incomplete nature of the acid digestion under these conditions allowed detection of the digest products along with the proteins from which they originated, which increased species identification confidence. The feasibility of this approach for the rapid identification of Bacillus species was further demonstrated by analyzing a mixture of B. subtilis str. 168 and B. anthracis str. Sterne spores.

Bioearosol mass spectrometry (BAMS) analyzes single particles in real time from ambient air, placing strict demands on instrument sensitivity. Modeling of the BAMS reflectron time of flight (TOF) with SIMION revealed design limitations associated with ion transmission and instrument sensitivity at higher masses. Design and implementation of a BAMS linear TOF with electrostatic ion guide and delayed extraction capabilities has greatly increased the sensitivity and mass range relative to the reflectron design. Initial experimental assessment of the new instrument design revealed improved sensitivity at high masses as illustrated when using standard particles of cytochrome C (m/z approximately 12,000), from which the compound's monomer, dimer (m/z approximately 24,000) and trimer (m/z approximately 36,000) were readily detected.

Bioaerosol mass spectrometry (BAMS) performs single-cell analysis in real time. However, the specificity of BAMS mass signatures has been limited by low sensitivity at high masses. To increase the mass range and sensitivity of BAMS, a novel design was developed that utilizes a linear flight tube with delayed extraction and an electrostatic ion guide. This study quantifies the sensitivity limits of the novel BAMS design and evaluates the feasibility of BAMS to detect higher mass biomarkers from single cells. All experiments were carried out using MALDI aerosol particles that were nebulized from solution. Sensitivity was assessed by generating particles with decreasing amounts of analyte via serial dilutions. The amount of analyte contained within each particle was calculated based on particle size, density, and molarity of the analyte within solution. A variety of biomolecular ions were studied and signals obtained from particles containing 300 zmol of maltopentaose, 132 zmol of alpha-cyclodextrin, and 14 zmol (approximately 8400 molecules) of gramicidin S are reported. The detection of 14 zmol of gramicidin S is to the best of our knowledge a record in sensitivity for MALDI TOF-MS.

Oligosaccharides are known to play important roles in many biological processes. In the study of oligosaccharides, collision-induced dissociation (CID) is the most common dissociation method to elucidate the sequence and connectivity. However, a disadvantage of CID is the decrease in both the degree and efficiency of dissociation with increasing mass. In the present study, we have successfully performed infrared multiphoton dissociation (IRMPD) on 39 O-linked mucin-type oligosaccharide alditols (both neutral and anionic). CID and IRMPD spectra of several oligosaccharides were also compared. They yielded nearly identical fragment ions corresponding to the lowest energy fragmentation pathways. The characteristic fragmentations of structural motifs, which can provide the linkage information, were similarly presented in both CID and IRMPD spectra. Multistage of CID (MS(3) or MS(4)) is commonly needed to completely sequence the oligosaccharides, while IRMPD of the same compounds yielded the fragment ions corresponding to the loss of the first residue to the last residue during a single-stage tandem MS (MS(2)). Finally, it is shown that the fragmentation efficiency of IRMPD increases with the increasing size of oligosaccharides.

Numerous American Indian tribal governments have introduced legalized gambling to enhance revenues. There have been notable financial successes as well as some confrontations with state governments and the exacerbation of factionalism on some reservations. The largest tribal entity in the United States, the Navajo, has not established a gaming industry. In 1994 the Navajo Tribal Council, following a veto of enabling legislation, referred the issue to the voters. Navajo voters rejected the referendum in the November, 1994 tribal general election. This article examines the Navajo electorate's rejection of gaming. Results varied by voting district (chapter). Exit polling conducted in a half-dozen chapters is used to analyze the effects of sex and age on the outcome and to describe the voters' reasons for their vote.

Sequence-specific polymers are the basis of the most promising approaches to bottom-up programmed assembly of nanoscale materials. Examples include artificial peptides and nucleic acids. Another class is oligo(N-functional glycine)s, also known as peptoids, which permit greater sidegroup diversity and conformational control, and can be easier to synthesize and purify. We have developed a set of peptoids that can be used to make inorganic nanoparticles more compatible with biological sequence-specific polymers so that they can be incorporated into nucleic acid or other biologically based nanostructures. Peptoids offer degrees of modularity, versatility, and predictability that equal or exceed other sequence-specific polymers, allowing for rational design of oligomers for a specific purpose. This degree of control will be essential to the development of arbitrarily designed nanoscale structures.

In recent years a major effort by several groups has been undertaken to identify bacteria by mass spectrometry at the single cell level. The intent of this review is to highlight the recent progress made in the application of single particle mass spectrometry to the analysis of microorganisms. A large portion of the review highlights improvements in the ionization and mass analysis of bio-aerosols, or particles that contain biologically relevant molecules such as peptides or proteins. While these are not direct applications to bacteria, the results have been central to a progression toward single cell mass spectrometry. Developments in single particle matrix-assisted laser desorption/ionization (MALDI) are summarized. Recent applications of aerosol laser desorption/ionization (LDI) to the analysis of single microorganisms are highlighted. Successful applications of off-line and on-the-fly aerosol MALDI to microorganism detection are discussed. Limitations to current approaches and necessary future achievements are also addressed.

Bioinformatic considerations are offered to illustrate strengths and limitations of the characterization of Bacillus spores based on proteomic interpretation of matrix-assisted laser desorption/ionization spectra. In particular, species-specific biomarkers are evaluated in the context of both experimental access and uniqueness in silico.

A proteomics approach is reported for the rapid recognition of genetically modified Escherichia coli bacteria. The approach targets a class of proteins required for genetic manipulation of bacteria with plasmids and alleviates the need to construct extensive libraries of toxins and other predicted payload proteins. Detection was performed using MALDI-TOF MS to monitor peptide products after an on-probe enzymatic digestion. Digestion products were identified by searching their postsource decay spectra using MASCOT. A 5 min digestion time was required to observe peptide products from the genetic insert as well as the host bacterium. This proteomics approach enables rapid detection of genetic manipulation along with information about the host organism, both of which have forensic applications.

We have fully characterized the mass spectral signatures of individual Bacillus atrophaeus spores obtained using matrix-free laser desorption/ionization bioaerosol mass spectrometry (BAMS). Mass spectra of spores grown in unlabeled, 13C-labeled, and 15N-labeled growth media were used to determine the number of carbon and nitrogen atoms associated with each mass peak observed in mass spectra from positive and negative ions. To determine the parent ion structure associated with fragment ion peaks, the fragmentation patterns of several chemical standards were independently determined. Our results confirm prior assignments of dipicolinic acid, amino acids, and calcium complex ions made in the spore mass spectra. The identities of several previously unidentified mass peaks, key to the recognition of Bacillus spores by BAMS, have also been revealed. Specifically, a set of fragment peaks in the negative polarity is shown to be consistent with the fragmentation pattern of purine nucleobase-containing compounds. The identity of m/z = +74, a marker peak that helps discriminate B. atrophaeus from Bacillus thuringiensis spores grown in rich media is [N1C4H12]+. A probable precursor molecule for the [N1C4H12]+ ion observed in spore spectra is trimethylglycine (+N(CH3)3CH2COOH), which produces a m/z = +74 peak when ionized in the presence of dipicolinic acid. A clear assignment of all the mass peaks in the spectra from bacterial spores, as presented in this work, establishes their relationship to the spore chemical composition and facilitates the evaluation of the robustness of "marker" peaks. This is especially relevant for peaks that have been used to discriminate Bacillus spore species, B. thuringiensis and B. atrophaeus, in our previous studies.

Single vegetative cells and spores of Bacillus atrophaeus, formerly Bacillus subtilis var. niger, were analyzed using bioaerosol mass spectrometry. Key biomarkers were identified from organisms grown in 13C and 15N isotopically enriched media. Spore spectra contain peaks from dicipolinate and amino acids. The results indicate that compounds observed in the spectra correspond to material from the spore's core and not the exosporium. Standard compounds and mixtures were analyzed for comparison. The biomarkers for vegetative cells were clearly different from those of the spores, consisting mainly of phosphate clusters and amino acid fragments.

Although electronic medical records (EMRs) are widely regarded as valuable tools in patient care, physicians in outpatient practices have been slow to adopt them. We sought to determine the current use of EMRs in area practices and identify physician attitudes related to their adoption.Fax and mail survey of randomly selected physician representatives of all outpatient practices of Internal Medicine (n=51) and Pediatrics (n=24) in Shelby County, Tenn. Scores on eight physician attitudes regarding barriers to EMR adoption were obtained using a Likert scale.Survey response rate was 55%, with 18% reporting current EMR use. This corresponds to an EMR penetration of 20% for Shelby County. Current users were significantly less likely (P=0.005) than non-users to feel that an EMR interferes with doctor-patient interaction and less likely (P=0.019) to have EMR privacy concerns. While differences noted in other attitudes did not reach statistical significance, a trend was seen toward EMR users being less concerned (P=.0502) about reliability of an EMR. Large practices were no more likely than smaller ones to be using an EMR. Internal Medicine and Pediatric participants responded similarly to all items. The number of years in practice had no demonstrable impact on physician responses to these survey items.In this West Tennessee physician population, EMR user and non-user attitudes markedly differed about impact on doctor-patient interaction and patient privacy. If such concerns could be addressed to the satisfaction of physicians considering EMRs in their practice, adoption rates might be increased.

The rapid chemical analysis of individual cells is an analytical capability that will profoundly impact many fields including bioaerosol detection for biodefense and cellular diagnostics for clinical medicine. This article describes a mass spectrometry-based analytical technique for the real-time and reagentless characterization of individual airborne cells without sample preparation. We characterize the mass spectral signature of individual Bacillus spores and demonstrate the ability to distinguish two Bacillus spore species, B. thuringiensis and B. atrophaeus, from one another very accurately and from the other biological and nonbiological background materials tested with no false positives at a sensitivity of 92%. This example demonstrates that the chemical differences between these two Bacillus spore species are consistently and easily detected within single cells in seconds.

Bioaerosol mass spectrometry is being developed to analyze and identify biological aerosols in real time. Characteristic mass spectra from individual bacterial endospores of Bacillus subtilis var. niger were obtained in a bipolar aerosol time-of-flight mass spectrometer using a pulsed 266-nm laser for molecular desorption and ionization. Spectra from single spores collected at an average fluence of approximately 0.1 J/cm2 frequently contain prominent peaks attributed to arginine, dipicolinic acid, and glutamic acid, but the shot-to-shot (spore-to-spore) variability in the data may make it difficult to consistently distinguish closely related Bacillus species with an automated routine. Fortunately, a study of the laser power dependence of the mass spectra reveals clear trends and a finite number of "spectral types" that span most of the variability. This, we will show, indicates that a significant fraction of the variability must be attributed to fluence variations in the profile of the laser beam.

The appearance of informative signals in the mass spectra of laser-ablated bio-aerosol particles depends on the effective ionization probabilities (EIP) of individual components during the laser ionization process. This study investigates how bio-aerosol chemical composition governs the EIP values of specific components and the overall features of the spectra from the bio-aerosol mass spectrometry (BAMS). EIP values were determined for a series of amino acid, dipicolinic acid, and peptide aerosol particles to determine what chemical features aid in ionization. The spectra of individual amino acids and dipicolinic acid, as well as mixtures, were examined for extent of fragmentation and the presence of molecular ion dimers, which are indicative of ionization conditions. Standard mixtures yielded information with respect to the significance of secondary ion plume reactions on observed spectra. A greater understanding of how these parameters affect EIP and spectra characteristics of bio-aerosols will aid in the intelligent selection of viable future biomarkers for the identification of bio-terrorism agents.

This book traces the history of the American Indians from their early mastery of the wilderness, through the classical era of their great civilizations, and the dark ages that followed European invasion and conquest. Jennings investigates how the Indians struggled to retain their cultural identity against efforts of government and society to force assimilation onto them. He documents the gradual population and cultural revival among Indians in the 20th century after their catastrophic decline in earlier years. By emphasizing the diversity of Indian cultures and the variety of their interactions with Europeans, Jennings refutes the legends characterizing the Indians as antagonistic warriors, and illuminates the complexity and significance of the realtionships that formed between the Indians and the settlers.

The recent growth of interest in Navajo archaeology suffers from the lack of an analytical foundation comparable to that which structures research on the prehistoric populations of the Southwest. Observations on the data potential of the ubiquitous sheep and goat corral are offered as a contribution to the effort to develop such a basis for Navajo archaeology. These corrals provide information on site function, site chronology, duration of site use, site abandonment and reuse, seasonality of site utilization, herd sizes, movements of residence groups, and economic behavior of the sites' inhabitants.

Over the last hundred years, the Navajo economy has changed from one based on animal husbandry, agriculture, arts and crafts to one in which the most significant returns derive from wage labor, welfare and Social Security. With the addition of these new sources, the contributions of women, men, and children in the economic sphere have changed substantially. This paper examines and enumerates the changes that have occurred in the Western Navajo community of Shonto.