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pathogenesis ( 4-6 ). The abundance and biological importance of the Mtb lipids has resulted in extensive and elegant studies to elucidate their structures and functions ( 1-3 ). In many cases, the lipids of Mtb are unique to this pathogen or shared only with other members of this genus.Earlier studies demonstrate variability in lipid profi les among different strains of Mtb ( 7-11 ) and that minor variations in the structure of individual lipids can occur with changes in the growth environment (12)(13)(14)(15)(16)(17)(18)(19)(20). However, targeted and nontargeted assays that monitor changes in Mtb lipid profi les are generally performed by traditional TLC-based methods ( 21 ), and global lipidomics analyses in Mtb have been restricted due to limits in the technology to detect and rapidly identify a large number of lipids in a single experiment. Two-dimensional NMR was recently applied to examine global mycobacterial lipid profi les, and this approach allowed for the identifi cation of key lipid differences in 13 C-enriched cellular extracts ( 22 ). Although this approach easily detects changes in lipid patterns, it is limited by the complexity of the NMR spectra and the overlapping chemical properties of many lipids. Alternatively, MS-based lipidomic strategies allowing simultaneous detection, identifi cation, and quantifi cation of structurally diverse lipid components of Mtb also were evaluated. Leavell and Leary ( 23 ) developed an algorithm to analyze high-resolution Fourier transform-ion cyclotron Abstract The cellular envelope of Mycobacterium tuberculosis is highly distinctive and harbors a wealth of unique lipids possessing diverse structural and biological properties. However, the ability to conduct global analyses on the full complement of M. tuberculosis lipids has been missing from the repertoire of tools applied to the study of this important pathogen. We have established methods to detect and identify lipids from all major M. tuberculosis lipid classes through LC/MS lipid profi ling. This methodology is based on efficient chromatographic separation and automated ion identifi cation through accurate mass determination and searching of a newly created database ( Mtb LipidDB) that contains 2,512 lipid entities. We demonstrate the sensitive detection of molecules representing all known classes of M. tuberculosis lipids from a single crude extract. We also demonstrate the ability of this methodology to identify changes in lipid content in response to cellular growth phases. This work provides a customizable framework and resource to facilitate future studies on mycobacterial lipid biosynthesis and

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Polyselenide anion speciation in aqueous solution has been investigated by electrospray mass spectrometry (ESMS). Two methods of control over this speciation are reported: countercation substitutions and pH adjustments. The countercation has been found to affect the degree of disproportionation or comproportionation. Solutions of sodium tetraselenide displayed numerous Se n 2and HSe n -species in neutral and basic soluions. In potassium solutions, the dominant species was Se 4 2-‚H 2 O. Solutions of cesium pentaselenide also showed preference for higher-order polyselenides. The pH dependence has been analyzed in terms of the percent Se n (n ) 2-5) present. Di-, tri-, tetra-, and pentaselenide species were present over the entire pH range investigated, and trends in selenide species versus pH were developed. For example, at pH > 9, tetraselenide species were dominant.

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The determination of sub-ppm concentrations of aqueous perfluoroalkylsulfonate (PFSt) anions, including perfluorooctylsulfonate (PFOS), has been accomplished with a relatively simple mass spectrometric procedure that does not require extraction of the analytes into an organic solvent or a chromatographic separation prior to injection into the negative-ion electrospray ionization mass spectrometer. Sample pretreatment was minimized and consisted of dilution of the aqueous samples of groundwater, surface water, tap water, and distilled water with acetonitrile, addition of dodecylsulfate (DDS) as an internal standard, and, in some cases, addition of known amounts of perfluorobutylsulfonate (PFBS) or PFOS for standard-addition experiments. The linear-response range for PFOS is 25.0 microg L(-1) to 2.5 mg L(-1). The lower limit of this range is three orders of magnitude lower than an equally straightforward chromatographic method. The relative errors for standard aqueous solutions containing only 25.0 microg L(-1) and 2.5 mg L(-1) PFOS are +/- 14% and +/- 7%, respectively, with 133 microg L(-1) DDS as the internal standard. The detection limit and quantification limit for PFOS in these standards are 5.0 microg L(-1) and 25.0 microg L(-1), respectively. Six different PFS anions, containing three to eight carbon atoms, were identified and quantified in an aqueous film-forming foam (AFFF) formulation using the method of standard additions. Two alkylsulfate anions and two perfluoroalkylcarboxylate anions were also identified in the AFFF formulation.

Monogalactosyldiacylglycerol (MGDG) was identified as a host recognition cue for larvae of the western corn rootworm Diabrotica virgifera virgifera LeConte. An active glycolipid fraction obtained from an extract of germinating maize roots was isolated with thin layer chromatography using a bioassay-driven approach. When analyzed with LC-MS (positive ion scanning), the assay-active spot was found to contain four different MGDG species: 18:3-18:3 (1,2-dilinolenoyl), 18:2-18:3 (1-linoleoyl, 2-linolenoyl), 18:2-18:2 (1,2-dilinoleoyl), and 18:2-16:0 (1-linoleoyl, 2-palmitoyl). A polar fraction was also needed for activity. When combined with a polar fraction containing a blend of sugars (glucose:fructose:sucrose:myoinositol), the isolated MGDG elicited a unique tight-turning behavior by neonate western corn rootworm larvae that is indicative of host recognition. In behavioral bioassays where disks treated with the active blend were exposed to successive sets of rootworm larvae, the activity of MGDG increased over four exposures, suggesting that larvae may be responding to compounds produced after enzymatic breakdown of MGDG. In subsequent tests with synthetic blends composed of theoretical MGDG-breakdown products, larval responses to four synthetic blends were not significantly different (P<0.5) than the response to isolated MGDG. GC-MS analysis showed modest increases in the amounts of the 16:0, 18:0, and 18:3 free fatty acids released from MGDG after a 30-min exposure to rootworm larvae, which is consistent with the enzymatic breakdown hypothesis.

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