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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 217 records · Page 12
All sourcesNASA NTRSDOE OSTI
DOE OSTI2016

Human serum albumin hydropersulfide is a potent reactive oxygen species scavenger in oxidative stress conditions such as chronic kidney disease

Recently, hydropersulfide (RSSH) was found to exist in mammalian tissues and fluids. Cysteine hydropersulfide can be found in free cysteine residues as well as in proteins, and it has potent antioxidative activity. Human serum albumin (HSA) is the most abundant protein in mammalian serum. HSA possesses a free thiol group in Cys-34 that could be a site for hydropersulfide formation. HSA hydropersulfide of high purity as a positive control was prepared by treatment of HSA with Na{sub 2}S. The presence of HSA hydropersulfide was confirmed by spectroscopy and ESI-TOFMS analysis where molecular weights of HSA hydropersulfide by increments of approximately 32 Da (Sulfur atom) were detected. The fluorescent probe results showed that Alexa Fluor 680 conjugated maleimide (Red-Mal) was a suitable assay and bromotrimethylammoniumbimane bromide appeared to be a selective reagent for hydropersulfide. The effect of oxidative stress related disease on the existence of albumin hydropersulfides was examined in rat 5/6 nephrectomy model of chronic kidney disease (CKD). Interestingly, the level of hydropersulfides in rat 5/6 nephrectomy model serum was decreased by a uremic toxin that increases oxidative stress in rat 5/6 nephrectomy model. Furthermore, we demonstrated that the levels of HSA hydropersulfide in human subjects were reduced in CKD but restored by hemodialysis using Red-Mal assay. We conclude that HSA hydropersulfide could potentially play an important role in biological anti-oxidative defense, and it is a promising diagnostic and therapeutic marker of oxidative diseases. - Highlights: • Hydropersulfide can behave as potent antioxidants. • We firstly detected human serum albumin hydropersulfide in healthy subjects. • Human serum albumin hydropersulfide in human subjects were reduced in chronic kidney disease but restored by hemodialysis.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2017

Exchange proteins directly activated by cAMP induce the proliferation of rat anterior pituitary GH3 cells via the activation of extracellular signal-regulated kinase

Cyclic adenosine 3′-5’-monophosphate (cAMP) plays a crucial role in regulating pituitary cell proliferation and hormone synthesis. Recent evidence suggests that exchange proteins directly activated by cAMP (Epacs) may mediate the effects of cAMP. Here we used rat anterior pituitary GH3 cells as the experiment model to demonstrate that forskolin increased the proliferation of GH3 cells and the phosphorylation of ERK1/2, and these effects were inhibited by PKA or Epac inhibitors. Epac activator 8-pCPT-2′-O-Me-cAMP increased GH3 cell proliferation and this was blocked by ESI-09, an Epac inhibitor. In contrast, forskolin-induced phosphorylation of CREB was unaffected by Epac inhibition. Notably, increased phosphorylation of ERK1/2 was correlated with increased cyclin D3 expression in GH3 cells. Furthermore, knockdown of Epac as well as B-Raf and MEK inhibitors blocked 8-pCPT-2′-O-Me-cAMP induced proliferation of GH3 cells and the phosphorylation of ERK1/2. In conclusion, our study suggests that Epac mediates cAMP induced pituitary cell proliferation via B-Raf and MAPK dependent mechanism. - Highlights: • PKA and Epac mediate cAMP induced GH3 cell proliferation. • Activation or knockdown of Epac stimulates or inhibits GH3 cell proliferation. • Epac induces the phosphorylation of ERK1/2 in GH3 cells via B-Raf. • B-Raf and MEK inhibitors block cAMP induced GH3 cell proliferation.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Laminin-1 induces endocytosis of 67KDa laminin receptor and protects Neuroscreen-1 cells against death induced by serum withdrawal

Highlights: • Laminin and YIGSR peptide induced internalization of 67KDa laminin receptor (67LR). • Agents that elevate intracellular cAMP also induced internalization of 67LR. • Adenylyl cyclase, protein kinase A, and Epac mediate the internalization of 67LR. • Early endosomes with 67LR and adenylyl cyclase induced sustained cell signaling. • Early endosome-mediated cell signaling protected neuronal cells from death. Although the function of laminin in the basement membrane is known, the function of soluble “neuronal” laminin is unknown. Since laminin is neuroprotective, we determined whether the soluble laminin-1 induces signaling for neuroprotection via its 67KDa laminin-1 receptor (67LR). Treatment of Neuroscreen-1 (NS-1) cells with laminin-1 or YIGSR peptide, which corresponds to a sequence in laminin-1 β1 chain that binds to 67LR, induced a decrease in the cell-surface expression of 67LR and caused its internalization. Furthermore, intracellular cAMP-elevating agents, dibutyryl-cAMP, forskolin, and rolipram, also induced this internalization. Both soluble laminin-1 and YIGSR induced a sustained elevation of intracellular cAMP under defined conditions, suggesting a causal role of cAMP in the endocytosis of 67LR. This endocytosis was not observed in cells deficient in protein kinase A (PKA) nor in cells treated with either SQ 22536, an inhibitor for adenylyl cyclase, or ESI-09, an inhibitor for the exchange protein directly activated by cAMP (Epac). In addition, when internalization occurred in NS-1 cells, 67LR and adenylyl cyclase were localized in early endosomes. Under conditions in which endocytosis had occurred, both laminin-1 and YIGSR protected NS-1 cells from cell death induced by serum withdrawal. However, under conditions in which endocytosis did not occur, neither laminin-1 nor YIGSR protected these cells. Conceivably, the binding of laminin-1 to 67LR causes initial signaling through PKA and Epac, which causes the internalization of 67LR, along with signaling enzymes, such as adenylyl cyclase, into early endosomes. This causes sustained signaling for protection against cell death induced by serum withdrawal.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Applications of mass spectrometry-based targeted and non-targeted lipidomics

Highlights: • Advances in mass spectrometry have expanded our knowledge of lipids. • Targeted and non-targeted lipidomics are powerful strategies with distinct features. • We briefly review the applications of these lipidomics strategies in lipid research. • Current limitations and challenges in lipidomics are also discussed. Recent advances in mass spectrometry have expanded our knowledge of lipids and lipid metabolic pathways involved in many (patho)physiological events. Targeted and non-targeted lipidomics are powerful analytical strategies with distinct features, and a combination of these two approaches is often employed to maximize the coverage of lipid species detected and quantified in complex biological matrices. This review briefly summarizes the applications of targeted and non-targeted lipidomics, mainly focusing on electrospray ionization–liquid chromatography–tandem mass spectrometry (ESI–LC–MS/MS), along with recent technical advances in the field. Current limitations and challenges in lipidomics and possible solutions are also discussed.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2019

Synthesis, characterization, anticancer evaluation and mechanisms of cytotoxic activity of novel 3-hydroxy-3-pyrrolin-2-ones bearing thenoyl fragment: DNA, BSA interactions and molecular docking study

In order to make a progress in discovering a new agents for chemotherapy with improved properties and bearing in mind the fact that substituted 3-hydroxy-3-pyrrolin-2-ones belong to a class of biologically active compounds, series of novel 1,5-diaryl-4-(2-thienylcarbonyl)-3-hydroxy-3-pyrrolin-2-ones were synthesized and characterized by spectral (UV–Vis, IR, NMR, ESI-MS), X-ray and elemental analysis. All compounds were examined for their cytotoxic effect on human cancer cell lines HeLa and MDA-MB 231 and normal fibroblasts (MRC-5). Four compounds, 3-hydroxy-1-(p-tolyl)-4-(2-thienylcarbonyl)-5-(4-chlorophenyl)-2,5-dihydro-1H-pyrrol-2-one (D10), 3-hydroxy-1-(3-nitrophenyl)-4-(2-thienylcarbonyl)-5-(4-(benzyloxy)phenyl)-2,5-dihydro-1H-pyrrol-2-one (D13), 3-hydroxy-1-(4-nitrophenyl)-4-(2-thienylcarbonyl)-5-(4-(benzyloxy)phenyl)-2,5-dihydro-1H-pyrrol-2-one (D14), and 3-hydroxy-1-(4-chlorophenyl)-4-(2-thienylcarbonyl)-5-(4-(benzyloxy)phenyl)-2,5-dihydro-1H-pyrrol-2-one (D15), that showed the highest cytotoxicity against malignant cells and the best selectivity towards normal cells were selected for further experiments. Results obtained by investigating mechanisms of cytotoxic activity suggest that selected 3-hydroxy-3-pyrrolin-2-one derivatives in HeLa cells induce apoptosis that is associated with S phase arrest (D13, D15, and D10) or unrelated to cell cycle distribution (D14). Additionally, to better understand their suitability for potential use as anticancer medicaments we studied the interactions between biomacromolecules (DNA or BSA) and D13 and D15. The results indicated that D13 and D15 have great affinity to displace EB from the EB-DNA complex through intercalation [K sv = (3.7 ± 0.1) and (3.4 ± 0.1) × 10 3 M -1 , respectively], an intercalative mode also confirmed through viscosity measurements. K a values, obtained as result of fluorescence titration of BSA with D13 and D15 [K a = (4.2 ± 0.2) and (2.6 ± 0.2) × 10 5 M, respectively], support the fact that a significant amount of the tested compounds could be transported and distributed through the cells. In addition, by DNA and BSA molecular docking study for D13, D14 and D15 is determined and predicted the binding mode and the interaction region. © 2019 Elsevier Inc.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2016

Interactions of cationic polystyrene nanoparticles with marine bivalve hemocytes in a physiological environment: Role of soluble hemolymph proteins

The bivalve Mytilus galloprovincialis has proven as a suitable model invertebrate for evaluating the potential impact of nanoparticles (NPs) in the marine environment. In particular, in mussels, the immune system represents a sensitive target for different types of NPs. In environmental conditions, both NP intrinsic properties and those of the receiving medium will affect particle behavior and consequent bioavailability/uptake/toxicity. However, the evaluation of the biological effects of NPs requires additional understanding of how, once within the organism, NPs interact at the molecular level with cells in a physiological environment. In mammalian systems, different NPs associate with serum soluble components, organized into a “protein corona”, which affects particle interactions with target cells. However, no information is available so far on the interactions of NPs with biological fluids of aquatic organisms. In this work, the influence of hemolymph serum (HS) on the in vitro effects of amino modified polystyrene NPs (PS-NH{sub 2}) on Mytilus hemocytes was investigated. Hemocytes were incubated with PS-NH{sub 2} suspensions in HS (1, 5 and 50 µg/mL) and the results were compared with those obtained in ASW medium. Cell functional parameters (lysosomal membrane stability, oxyradical production, phagocytosis) were evaluated, and morphological changes were investigated by TEM. The activation state of the signalling components involved in Mytilus immune response (p38 MAPK and PKC) was determined. The results show that in the presence of HS, PS-NH{sub 2} increased cellular damage and ROS production with respect to ASW medium. The effects were apparently mediated by disregulation of p38 MAPK signalling. The formation of a PS-NH{sub 2}-protein corona in HS was investigated by centrifugation, and 1D- gel electrophoresis and nano-HPLC-ESI-MS/MS. The results identified the Putative C1q domain containing protein (MgC1q6) as the only component of the PS-NH{sub 2} hard protein corona in Mytilus hemolymph. These data represent the first evidence for the formation of a NP bio-corona in aquatic organisms and underline the importance of the recognizable biological identity of NPs in physiological exposure medium when testing their potential impact environmental model organisms. Although the results obtained in vitro do not entirely reflect a realistic exposure scenario and the more complex formation of a bio-corona that is likely to occur in vivo, these data will contribute to a better understanding of the effects of NPs in marine invertebrates. - Highlights: • The effects of PS-NH2 NPs on Mytilus hemocytes were compared in serum-HS and ASW. • HS increased cellular/lysosomal damage, ROS production and p-p38MAPK levels. • NP-corona proteins in HS were isolated and identified by MS. • NP-protein coronas in biological fluids can affect NP impact in marine species.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2018

Perfluoroalkyl substances, bone density, and cardio-metabolic risk factors in obese 8–12 year old children: A pilot study

Highlights: • Cross sectional study of 48 obese children aged 8–12 years. • Serum Perfluoroalkyl substances (PFASs), measured by UPLC-ESI-MS/MS. • In 23 children, bone health assessed using calcaneal quantitative ultrasound. • PFNA showed significant negative relationship with bone health. • PFNA associated with raised systolic blood pressure and lipids. Perfluoroalkyl substances (PFASs), including perfluorooctanoic acid (PFOA), perfluorooctane sulfonic acid (PFOS), perfluorohexane sulfonic acid (PFHxS), and perfluorononanoic acid (PFNA), have been associated with adverse bone, and metabolic changes in adults. However association of PFASs with bone health in children is understudied. Considering their role as endocrine disruptors, we examined relationships of four PFASs with bone health in children.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2018

Occurrence and ecological risk of pharmaceuticals in river surface water of Bangladesh

Highlights: • Twelve pharmaceuticals were investigated in river surface water. • Nine pharmaceuticals were detected at ng L{sup −1} level. • MNZ was dominant followed by TMP, ERY-H{sub 2}O and SMX. • Contamination of SMX, TMP and TYL was associated with fed aquaculture. • Preliminary ecological risk assessment showed the medium to low risks. Pharmaceutical contamination in the aquatic environment is a global issue that affects aquatic animals, micro-organisms and human health. The occurrence and preliminary ecological risk of 12 (11 antibiotics and 1 antiepileptic drug) pharmaceuticals were investigated for the first time in the surface water of the old Brahmaputra River, where open-water-fed aquaculture activities are being practiced in Bangladesh. The pharmaceuticals were quantified by high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS), operated with positive electrospray ionization (ESI{sup +}) and a multiple reaction monitoring (MRM) mode. Nine pharmaceuticals were detected in the river surface water, whereas three were below the limit of detection (LOD). Metronidazole was detected in all the samples with concentrations ranging from 0.05 to 13.51 ng L{sup −1}. Trimethoprim had the second highest frequency of detection (95%) with the highest concentration (17.20 ng L{sup −1}). The ranges of concentration and detection frequency of sulfonamides and macrolides were < LOD-11.35 and < LOD-16.68 ng L{sup −1}; 35–70 and 60–85%, respectively, whereas carbamazepine was in the range of < LOD-8.80 ng L{sup −1} and had a detection frequency of 65%. The concentrations of sulfamethoxazole, trimethoprim, erythromycin-H{sub 2}O and tylosin were distinctly higher in the fed aquaculture areas. The principal component analysis confirmed that fed aquaculture activities contributed most of the pharmaceutical contamination in the river surface water. Hospitals, nursing homes, sewage wastewater or surface runoff from the surrounding areas might all contribute to the presence of metronidazole and carbamazepine. The preliminary ecological risk assessment revealed that sulfamethoxazole, erythromycin-H{sub 2}O and tylosin showed medium risk, and carbamazepine displayed low risk to sensitive aquatic organisms for maximum measured concentrations. Thus, this study suggests that pharmaceutical contamination in different rivers and seasons needs to be quantified, and ecological as well as human health risks need to be assessed in Bangladesh.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2018

Bisphenol-A in breast adipose tissue of breast cancer cases and controls

Highlights: • BPA was detected in breast adipose tissue of breast cancer cases and controls. • Mean BPA concentrations were similar between cases and controls. • BPA concentrations were highly variable within- and between-breasts from a single patient. We evaluated whether bisphenol-A (BPA) could be quantified in breast adipose tissue samples provided by 36 breast cancer mastectomy patients and 14 reduction mammoplasty patients. Samples of breast adipose tissue were collected and BPA concentration was quantified using HPLC-ESI-MS/MS. BPA was detectable above the limit of quantitation of 0.38 ng/g in 30.6% of samples. BPA concentrations varied within- and between breasts and were similar between cases and controls (0.39 vs 0.41 ng/g, p = 0.74).

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2018

Comparative estrogenicity of endogenous, environmental and dietary estrogens in pregnant women I: Serum levels, variability and the basis for urinary biomonitoring of serum estrogenicity

Biomonitoring of human exposure to estrogens most frequently focuses on environmental and dietary estrogens, and infrequently includes measures of exposure to potent endogenous estrogens present in serum. Pregnancy is a developmentally sensitive period during which “added” serum estrogenicity exceeding normal intra-individual daily variability may be of particular relevance. Here, we made repeated measurements of serum concentrations of estrone (E1), estradiol (E2), estriol (E3), estetrol (E4), daidzein (DDZ), genistein (GEN) and bisphenol A (BPA) in thirty pregnant women using ultra-performance liquid chromatography coupled with tandem mass spectrometry detection (UPLC-MS/MS) and electrospray ionization (ESI). Serum E1, E2, and E3 concentrations varied significantly (coefficients of variation 9–10%) with broad ranges across the cohort: 1.61–85.1 nM, 9.09–69.7 nM, and 1.5–36.3 nM respectively. BPA (undetected, estimated from total exposure), DDZ and GEN concentrations were 1-5 orders of magnitude lower. The 24-h urinary elimination profiles of endogenous estrogens were each strongly correlated with their corresponding serum concentrations (Pearson's Correlation Coefficients of 0.83 (E1), 0.84 (E2) and 0.94 (E3)). Lastly, a multivariate regression analysis produced equations for estimating serum concentrations of E1, E2, E3, E4, GEN and DDZ from urinary elimination rates and gestation period, an important step towards non-invasive biomonitoring for assessment of “added” estrogenicity during pregnancy.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2016

Photochemical behavior of the quadruply metal-metal bonded [Tc 2 Cl 8 ] 2– anion in acetonitrile

Here, the photochemical behavior of [Tc 2 Cl 8 ] 2– was investigated in acetonitrile. The speciation of Tc before and after irradiation at 254 nm was performed by UV-visible spectroscopy and electrospray ionization mass spectrometry (ESI-MS). Upon irradiation at 254 nm, [Tc 2 Cl 8 ] 2– was unstable, the scission of the Tc ≡ Tc unit occurred and the complex [TcCl 4 (CH 3 CN) 2 ] was identified. The disappearance rate of [M 2 Cl 8 ] 2– (M = Tc, Re) under irradiation has been measured and was ~7.5 time faster for Tc than for Re.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2013

Influence of heteroanion and ammonium cation size on the composition and gas-phase fragmentation of polyoxovanadates

This paper describes the results of a systematic experimental investigation of the influence of different size cationic ammonium ligands and heteroanions on the composition, ionic charge state and gas-phase fragmentation pathways of anionic polyoxovanadates synthesized in solution. Four separate solutions of olyoxometalates (POMs) were prepared using all possible combinations of the tetraethylammonium [(C2H5)4N+] ligand, chloride (Cl-) heteroanion, tetrabutylammonium [(C4H9)4N+] ligand and acetate (CH3CO2-) heteroanion. Employing electrospray ionization combined with high-resolution mass spectrometry (ESI-MS) we demonstrate that POM solutions synthesized using the small [(C2H5)4N+] ligand and Cl-heteroanion are composed predominately of large doubly and triply charged chlorine containing clusters with a size distribution centered at fourteen vanadium atoms. POM solutions prepared using the Cl- anion and [(C4H9)4N+] ligand are shown to contain slightly larger clusters with fifteen and sixteen vanadium atoms, thereby indicating that the size of the cationic ammonium ligand exerts only a weak influence on the polymerization of polyoxovanadates. POM solutions prepared using (C2H5)4NCl and (C4H9)4NCl also produced peaks consistent with the attachment of one and two ammonium cations to the larger clusters. Solutions prepared using the large CH3CO2 - heteroanion, in contrast, are demonstrated to contain much smaller singly and doubly *Manuscript Click here to view linked References 2 charged clusters with a size distribution centered at six vanadium atoms. In addition, while incorporation of one and two ammonium ligands into the smaller clusters was observed, no POMs containing the CH3CO2 - heteroanion were identified. The gas-phase fragmentation pathways of representative POMs containing one and two ammonium ligands were examined using collision induced dissociation (CID) and mass spectrometry. Similar primary fragmentation pathways involving partial loss of a ligand -[(CxHy)3N+ x = 2,4; y = 5,9] were observed for clusters containing both one and two ligands largely independent of the size, composition and charge state of the precursor ion. The [(C4H9)4N+] ligand was found to exhibit stronger interactions with the core of the POMs resulting in higher abundances of fragment ions containing (C4H9) units compared to (C2H5) units from [(C2H5)4N+]. These results provide fundamental insight into the interactions between anionic metal oxide clusters, heteroanions and cationic ammonium ligands that are responsible for the size and composition controlled synthesis of POMs in solution.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2022

Dehydration of gas-phase benzyl amine alcohols studied at atmospheric pressure

Gas-phase ion chemistry is a useful approach for the investigation of physical organic chemistry and the study of reactivity, structure, and thermochemical properties of ionic and neutral organic substrates. Herein, one-step dehydration of gas-phase benzyl amine alcohols to give benzazetidines was discovered without the use of catalysts. Mechanistic investigations of the gas-phase dehydration reactions were explored to study the possible influence of charged microdroplet acceleration during electrospray ionization (ESI) and collisions occurring during ion transfer through the atmospheric pressure interface (API) of the mass spectrometer. The product ion distribution was observed to be less sensitive to droplet effects (spray distance and voltage), and temperature of transfer capillary in the API. However, the product ion distribution exhibit high sensitivity to different S-lens radio frequency (RF) voltage of the API, providing evidence that the dehydration reaction is driven by collisions during ion transfer in the API. With this insight, we developed atmospheric pressure thermal dissociation platform that allowed thermal-induced collisions outside of the mass spectrometer when the benzyl amine alcohols are carried through a heated coiled tube. Up to 98% dissociation efficiency was achieved with the coiled tube heated to a temperature of 200 °C, yielding only the desired benzazetidine product. Finally, by performing the atmospheric pressure thermal dissociation experiment outside of the mass spectrometer, we believe it will be straightforward to collect the dehydration product.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2023

Developing a peptide concatemer (PepCon) as a process control for LC-MS based proteomics

Quantitative concatenated peptide (QconCAT) technology is currently used for the absolute quantification of proteins of interest in a biological sample. It relies on artificially created proteins that are concatenations of different, isotopically labeled peptides. Because these isotopically labeled peptides are at a 1:1 ratio and correspond to naturally occurring peptides in the biological sample, each peptide can be used as a standard for the absolute quantitation of all proteins of interest at once. Although the QconCAT technology has utility for quantitation of known peptides in a mixture, it is not helpful for scientists who need a proteomics standard (1) that can be spiked into a protein mixture at an extremely low level, (2) that can be co-purified during sample fractionation, and (3) that is optimized for ESI used in mass spectrometry. Thus, there exists a need for an ideal standard protein that is large enough to behave as a protein but consists of multiple, concatenated copies of the same peptide, which, upon digestion, amplifies (e.g., >10-fold) into a detectable peptide species. Here, this technical report describes the development of “PepCon”, a peptide concatemer that fulfills this unmet need.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2017

Recent advances in the detection of specific natural organic compounds as carriers for radionuclides in soil and water environments, with examples of radioiodine and plutonium

Among the key environmental factors influencing the fate and transport of radionuclides in the environment is natural organic matter (NOM). While this has been known for decades, there still remains great uncertainty in predicting NOM-radionuclide interactions because of lack of understanding of radionuclide interactions with the specific organic moieties within NOM. Furthermore, radionuclide-NOM studies conducted using modelled organic compounds or elevated radionuclide concentrations provide compromised information related to true environmental conditions. Thus, sensitive techniques are required not only for the detection of radionuclides, and their different species, at ambient and/or far-field concentrations, but also for potential trace organic compounds that are chemically binding these radionuclides. GC-MS and AMS techniques developed in our lab are reviewed in this paper that aim to assess how two radionuclides, iodine and plutonium, form strong bonds with NOM by entirely different mechanisms; iodine tends to bind to aromatic functionalities, whereas plutonium binds to N-containing hydroxamate siderophores at ambient concentrations. While low-level measurements are a prerequisite for assessing iodine and plutonium migration at nuclear waste sites and as environmental tracers, it is necessary to determine their in-situ speciation, which ultimately controls their mobility and transport in natural environments. Finally and more importantly, advanced molecular-level instrumentation (e.g., nuclear magnetic resonance (NMR) and Fourier-transform ion cyclotron resonance coupled with electrospray ionization (ESI-FTICRMS) were applied to resolve either directly or indirectly the molecular environments in which the radionuclides are associated with the NOM.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2023

A Bis(imidazole)-based cysteine labeling tool for metalloprotein assembly

Precise metal-protein coordination by design remains a considerable challenge. Polydentate, high-metal-affinity protein modifications, both chemical and recombinant, can enable metal localization. However, these constructs are often bulky, conformationally and stereochemically ill-defined, or coordinately saturated. Here, we expand the biomolecular metal-coordination toolbox with the irreversible attachment to cysteine of bis(1-methylimidazol-2-yl)ethene (“BMIE”), which generates a compact imidazole-based metal-coordinating ligand. Conjugate additions of small-molecule thiols (thiocresol and N-Boc-Cys) with BMIE confirm general thiol reactivity. The BMIE adducts are shown to complex the divalent metal ions Cu ++ and Zn ++ in bidentate (N 2 ) and tridentate (N 2 S*) coordination geometries. Cysteine-targeted BMIE modification (>90% yield at pH 8.0) of a model protein, the S203C variant of carboxypeptidase G2 (CPG2), measured with ESI-MS, confirms its utility as a site-selective bioconjugation method. ICP-MS analysis confirms mono-metallation of the BMIE-modified CPG2 protein with Zn ++ , Cu ++ , and Co ++ . EPR characterization of the BMIE-modified CPG2 protein reveals the structural details of the site selective 1:1 BMIE-Cu ++ coordination and symmetric tetragonal geometry under physiological conditions and in the presence of various competing and exchangeable ligands (H 2 O/HO – , tris, and phenanthroline). An X-ray protein crystal structure of BMIE-modified CPG2-S203C demonstrates that the BMIE modification is minimally disruptive to the overall protein structure, including the carboxypeptidase active sites, although Zn ++ metalation could not be conclusively discerned at the resolution obtained. The carboxypeptidase catalytic activity of BMIE-modified CPG2-S203C was also assayed and found to be minimally affected. Finally, these features, combined with ease of attachment, define the new BMIE-based ligation as a versatile metalloprotein design tool, and enable future catalytic and structural applications.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2022

Analysis of phospholipids and triacylglycerols in intravenous lipid emulsions

Intravenous lipid emulsions (ILEs) are used for parenteral nutrition, providing a vital source of essential fatty acids and concentrated energy for patients who are unable to absorb nutrients via the digestive track. They are commonly used to treat local and non-local anesthetic toxicity, and lipophilic drug overdose. ILE are composed of natural lipids, and the composition of these natural lipids can be varied based on their source. The lipids are susceptible to hydrolytic degradation with time, resulting various lipid degradation products such as Lysophosphatidylcholines (LPs), affecting the actual composition of nutrients in the formulation. As a result, the identification and quantification of lipid components, including degradation products, in ILEs are crucial in quality control. In this study, lipids from different batches of ILE Intralipid® 20%, were separated and identified using a UHPLC-ESI-QTOF system and SimLipid® high throughput lipid identification software. Out of 47 lipids identified, 34 were phospholipids (PLs) and the others were triacylglycerols (TAGs). Most of the phospholipids detected were phosphatidylcholines (PC) and Lysophosphatidylcholines (LPC). A total of 9 LPCs, 18 PCs, 6 phosphoethanolamines (PEs), and 1 sphingomyelin (SM) were identified. The LPCs concentration changed with the manufacturing date and storage time. Furthermore, this UHPLC method enabled the identification and quantification of lipids and their decomposition products in complex ILE emulsion mixtures on a single 20-minute chromatographic run.

60 APPLIED LIFE SCIENCES↗