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We present a new wide-ranging correlation for the viscosity of nitrogen based on critically evaluated experimental data as well as ab-initio calculations. The correlation is designed to be used with densities from an existing equation of state, which is valid from the triple point to 1000 K, at pressures up to 2200 MPa. The estimated uncertainty (at the 95% confidence level) for the viscosity varies depending on the temperature and pressure, from a low of 0.2% in the dilute-gas range near room temperature, to 4% for the liquid phase at pressures from saturation up to 34 MPa, and maximum of 8% in the supercritical region at pressures above 650 MPa. Extensive comparisons with experimental data are provided., Supplementary Information: The online version contains supplementary material available at 10.1007/s10765-024-03440-1., Competing Interests: Competing InterestsThe authors declare no competing interests., (© This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply 2024.)

Vapor pressure ( p sat ) data are needed to assess the potential use of terpenes as breath markers of recent cannabis use. Herein, a recently introduced gas-saturation method for p sat measurements, known as dynamic vapor microextraction (DVME), was used to measure p sat for the terpene (±)-3,7-dimethylocta-1,6-dien-3-ol, commonly known as linalool. The DVME apparatus utilizes inexpensive and commercially available components, a low internal volume, and helium carrier gas to minimize nonideal mixture behavior. In the temperature range from 314 K to 354 K, DVME-based measurements of the p sat of linalool ranged from 81 Pa to 1250 Pa. With a measurement period of 30 min, the combined standard uncertainty of these measurements ranged from 0.0358· p sat to 0.0584· p sat , depending on temperature. The DVME-based measurements agree with a Wagner correlation of available literature data. We demonstrate that DVME produces accurate results for values of p sat that are 200 times higher than in the DVME validation study with n-eicosane (C 20 H 42 ). The oxidative stability of linalool was improved by the addition of 0.2 mass % of the antioxidant tert -butylhydroquinone., Competing Interests: The authors declare no competing financial interest.

We compiled an experimental database for the surface tension of binary mixtures containing a wide variety of fluids, from the chemical classes (water, alcohols, amines, ketones, linear and branched alkanes, naphthenes, aromatics, refrigerants, and cryogens). The resulting data set includes 65 pure fluids and 154 binary pairs with a total of 8205 points. We used this database to test the performance of a parachor model for the surface tension of binary mixtures. The model uses published correlations to determine the parachors of the pure fluids. The model has a single, constant binary interaction parameter for each pair that was found by fitting experimental mixture data. It can be also used in a predictive mode when the interaction parameters are set to zero. We present detailed comparisons on the performance of the model for both cases. In general, the parachor model in a predictive mode without fitted interaction parameters can predict the surface tension of binary mixtures of non-polar mixtures such as linear and branched alkanes, linear and branched alkanes with naphthenes, aromatics with aromatics, aromatics with naphthenes, and mixtures of linear alkanes of similar sizes with an average absolute percentage deviation of about 3 % or less. Polar mixtures of halocarbons with other halocarbons and also polar/nonpolar mixtures of alkanes with halocarbons could be modeled with an average absolute deviation of less than 0.35 mN·m -1 with the use of a binary interaction parameter. The parachor model even with a fitted binary interaction parameter performs poorly for mixtures of water and organic compounds and is not recommended., Supplementary Information: The online version contains supplementary material available at 10.1007/s10765-023-03216-z., Competing Interests: Conflict of interestThe authors have no conflicts to declare., (© The Author(s) 2023.)

The NIST REFPROP software program is a powerful tool for calculating thermophysical properties of industrially important fluids, and this manuscript describes the models implemented in, and features of, this software. REFPROP implements the most accurate models available for selected pure fluids and their mixtures that are valid over the entire fluid range including gas, liquid, and supercritical states, with the goal of uncertainties approaching the level of the underlying experimental data. The equations of state for thermodynamic properties are primarily of the Helmholtz energy form; a variety of models are implemented for the transport properties. We document the models for the 147 fluids included in the current version. A graphical user interface generates tables and provides extensive plotting capabilities. Properties can also be accessed through third-party apps or user-written code via the core property subroutines compiled into a shared library. REFPROP disseminates international standards in both the natural gas and refrigeration industries, as well as standards for water/steam., Competing Interests: The authors declare no competing financial interest., (Not subject to U.S. Copyright. Published 2022 by American Chemical Society.)

A total of 2160 thermal conductivity data points, measured using a transient hot-wire instrument, are reported for binary mixtures of R-134a, R-1234yf, and R-1234ze(E) refrigerants from 200 to 340 K to pressures of 12 MPa for mixtures containing R-1234yf and to 50 MPa for R-134a/1234ze(E) mixtures. Data are reported at compositions of approximately (0.33/0.67) mole fraction and (0.67/0.33) mole fraction for each binary mixture investigated. The estimated relative expanded uncertainty of the thermal conductivity measurements is less than 2%. The data are used to refit binary interaction parameters for the Extended Corresponding States (ECS) model implemented in REFPROP (version 10.0). Additionally, the data in this study are used to assess the performance of a generalized entropy scaling model for refrigerants. Finally, the strengths and weaknesses of the ECS and entropy scaling models are compared., Competing Interests: The authors declare no competing financial interest. Commercial equipment, instruments, or materials are identified only to adequately specify certain procedures. Such identification does not imply recommendation or endorsement by the National Institute of Standards and Technology, nor does it imply that the identified products are necessarily the best available for the purpose.

We present a new wide-ranging correlation for the viscosity of ethane-1,2-diol (ethylene glycol) based on critically evaluated experimental data. The correlation is designed to be used with an existing equation of state, and it is valid from the triple point to 465 K, at pressures up to 100 MPa. The estimated uncertainty is 4.9 % (at the 95 % confidence level), except in the dilute-gas region which is estimated to be 15 %, as there are no measurements in this region for comparison. The correlation behaves in a physically reasonable manner when extrapolated to 750 K and 250 MPa, however care should be taken when using the correlations outside of the validated range.

A new wide-ranging correlation for the viscosity of xenon, based on the most recent theoretical calculations and critically evaluated experimental data, is presented. The correlation is designed to be used with an existing equation of state, and it is valid from the triple point to 750 K, at pressures up to 86 MPa. The estimated expanded uncertainty (at a coverage factor of k = 2) varies depending on the temperature and pressure, from 0.2 % to 3.6 %. A term accounting for the critical enhancement is also included. The correlation behaves in a physically reasonable manner when extrapolated to 200 MPa, however care should be taken when using the correlations outside of the validated range.

We present a new wide-ranging correlation for the thermal conductivity of ethane-1,2-diol (ethylene glycol) based on critically evaluated experimental data. The correlation is designed to be used with an existing equation of state, and it is valid from the triple point to 475 K, at pressures up to 100 MPa. The estimated uncertainty is 2.2 % (at the 95 % confidence level), except in the dilute-gas region which is estimated to be 20 %, as there are no measurements in this region for comparison.

As we enter the "fourth generation" of refrigerants, we consider the evolution of refrigerant molecules, the ever-changing constraints and regulations that have driven the need to consider new molecules, and the advancements in the tools and property models used to identify new molecules and design equipment using them. These separate aspects are intimately intertwined and have been in more-or-less continuous development since the earliest days of mechanical refrigeration, even if sometimes out-of-sight of the mainstream refrigeration industry. We highlight three separate, comprehensive searches for new refrigerants-in the 1920s, the 1980s, and the 2010s-that sometimes identified new molecules, but more often, validated alternatives already under consideration. A recurrent theme is that there is little that is truly new. Most of the "new" refrigerants, from R-12 in the 1930s to R-1234yf in the early 2000s, were reported in the chemical literature decades before they were considered as refrigerants. The search for new refrigerants continued through the 1990s even as the hydrofluorocarbons (HFCs) were becoming the dominant refrigerants in commercial use. This included a return to several long-known natural refrigerants. Finally, we review the evolution of the NIST REFPROP database for the calculation of refrigerant properties., Competing Interests: The authors declare no competing financial interest.

New experimental data for the thermal conductivity of cis -1,1,1,4,4,4-hexafluoro-2-butene (R-1336mzz(Z)) are reported for vapor, liquid and supercritical states. These data were obtained with transient hot-wire apparatus over the temperature range from 192 K to 498 K and at pressures from 0.05 MPa to 69 MPa. These data were used to develop a wide-range correlation for the thermal conductivity of the vapor, liquid, and supercritical fluid. The experimental data reported here have an uncertainty of 1 % for the liquid and supercritical regions (densities above 600 kg·m -3 ), 1.5 % for vapor and supercritical regions (pressures greater than or equal to 1 MPa and densities less than 200 kg·m -3 ), 3 % for supercritical states (densities between 200 kg·m -3 and 600 kg·m -3 ), and 3 % for vapor and supercritical states (pressures below 1 MPa). The thermal-conductivity correlation developed in this work is estimated to have an expanded relative uncertainty, at a 95 % confidence level, ranging from approximately 1.4 % to 4.2 % depending on the temperature and pressure, with larger uncertainties in the critical region., Competing Interests: Declarations The authors declare no competing financial interest.

By building on the Advanced Distillation Curve (ADC) approach to measuring the volatility of fuels and other fluid mixtures, the ADC with Reflux or ADCR technique was developed to address the difficulty of experimentally determining the vapor-liquid equilibrium of fluids containing many components. For fuels and other multicomponent mixtures, the ADCR collects data about the chemical compositions of both liquid and vapor phases across a range of temperatures, elucidating the two-phase region at constant pressure. Two simple mixtures were used to demonstrate the ADCR method: an n -decane/ n -tetradecane binary and the Huber-Bruno surrogate, a ternary mixture designed to represent the volatility of an aviation turbine kerosene. These mixtures were chosen to test the method because they have been extensively studied and modeled in previous work. For both test fluids, the ADCR measurements of vapor-liquid equilibrium were in good agreement with model predictions. We conclude that the ADCR is a useful method for determining the T-P-x-y behavior of fluid mixtures with many components. The experimental approach presented may support the development of fuels, design of separations, and forensic sciences that use vapor analysis, especially arson fire debris analysis, by providing quantitative data with well-characterized uncertainty describing the relationships between the vapor and condensed phases of a fuel subjected to thermal weathering., Competing Interests: The authors declare no competing financial interest., (Copyright © 2019 American Chemical Society.)

The available experimental data for the thermal conductivity of 13 inorganic molten salts have been critically examined with the intention of establishing thermal conductivity reference correlations. All experimental data have been categorized into primary and secondary data according to the quality of measurement specified by a series of criteria. Standard reference correlations are proposed for the following molten salts (with estimated uncertainties at the 95 % confidence level given in parentheses): LiNO 3 (7 %), NaNO 3 (7 %), KNO 3 (15 %), NaBr (15 %), KBr (15 %), RbBr (15 %), LiCl (17 %), NaCl (20 %), KCl (17 %), RbCl (17%), CsCl (10 %), NaI (17 %), and RbI (20 %).

In this paper, reference values and reference correlations for the thermal conductivity and viscosity of pure fluids are reviewed. Reference values and correlations for the thermal conductivity and the viscosity of pure fluids provide thoroughly evaluated data or functional forms and serve to help calibrate instruments, validate or extend models, and underpin some commercial transactions or designs, among other purposes. The criteria employed for the selection of thermal conductivity and viscosity reference values are also discussed; such values, which have the lowest uncertainties currently achievable, are typically adopted and promulgated by international bodies. Similar criteria are employed in the selection of reference correlations, which cover a wide range of conditions, and are often characterized by low uncertainties in their ranges of definition.

New experimental data for the thermal conductivity of 1,1,1,2,2,4,5,5,5-nonafluoro-4-(trifluoromethyl)-3-pentanone (Novec 649) are reported for vapor, liquid and supercritical states. These new experimental data were obtained with transient hot-wire apparatus over the temperature range from 183 K to 501 K and at pressures from 0.02 MPa to 69 MPa. These data were used to develop a wide-range correlation for the thermal conductivity of the vapor, liquid and supercritical fluid. The experimental data reported here have an uncertainty of 1 % for the liquid and supercritical regions (densities > 600 kg m -3 ), 1.5 % for vapor and supercritical regions (pressures ≥ 1 MPa and densities < 200 kg m -3 ), 3 % for supercritical states (200 kg m -3 ≤ densities ≤ 600 kg m -3 ), and 3 % for vapor and supercritical states (pressures < 1 MPa). The thermal-conductivity correlation developed in this work is estimated to have an expanded relative uncertainty, at a 95 % confidence level, ranging from approximately 1 % to 4 % that depends upon the temperature and pressure, with larger uncertainties in the critical region., Competing Interests: The authors declare no competing financial interest.

This paper presents a new wide-ranging correlation for the thermal conductivity of n -hexadecane based on critically evaluated experimental data. The correlation is designed to be used with a recently published equation of state, and it is valid from the triple point up to 700 K and pressures up to 50 MPa. We estimate the uncertainty at a 95% confidence level to be 4% over the aforementioned range, with the exception of the dilute-gas range where the uncertainty is 2.7% over the temperature range 583 to 654 K. The correlation behaves in a physically reasonable manner when extrapolated to the full range of the equation of state, but the uncertainties are larger outside of the validated range, and also in the critical region.

This paper presents a new wide-ranging correlation for the viscosity of ammonia based on critically evaluated experimental data. The correlation is designed to be used with a recently developed equation of state, and it is valid from the triple point to 725 K, at pressures up to 50 MPa. The estimated uncertainty varies depending on the temperature and pressure, from 0.6% to 5%. The correlation behaves in a physically reasonable manner when extrapolated to 100 MPa, however care should be taken when using the correlations outside of the validated range.

The Paris Agreement on climate change, in which many nations have agreed to limit greenhouse gas emissions, has spurred interest in developing working fluids with low global warming potential (GWP) that can satisfy environmental concerns and have thermophysical properties that can meet engineering performance requirements. One such fluid is 1,1,1,2,2,4,5,5,5-nonafluoro-4-(trifluoromethyl)-3-pentanone (also known as Novec-649 and Novec-1230), which has potential applications in organic Rankine cycles (ORC), electronics cooling, computer/data center cooling and fire extinguishing. In this work, viscosity measurements of 1,1,1,2,2,4,5,5,5-nonafluoro-4-(trifluoromethyl)-3-pentanone were reported. The measurements were performed over the temperature range of (243 to 373) K and at pressures up to 40 MPa using a vibrating-wire viscometer. The combined expanded uncertainty of the reported viscosity was 2 % with a confidence level of 0.95 ( k = 2). These experimental data were used to develop a viscosity correlation that covers a wide temperature and pressure range, with an estimated uncertainty at a 95% confidence level of 2% for the liquid phase from (240 to 400) K at pressures up to 40 MPa., Competing Interests: Notes The authors declare no competing financial interest.

New experimental data on the thermal conductivity of trans -1-chloro-3,3,3-trifluoropropene (R1233zd(E)) are reported that allow the development of wide-range correlations. These new experimental data, covering a temperature range of 204 K to 453 K at pressures from 0.1 MPa to 67 MPa, are used to develop a correlation for the thermal conductivity. The experimental data reported here have an uncertainty of (1 - 1.5) % for liquid measurements and for gas at pressures above 1 MPa, increasing to (3 - 4) % for gas at low pressures (less than 1 MPa) and near the gas-liquid critical point. Based on the uncertainty of and comparisons with the present data, the thermal-conductivity correlation for R1233zd(E) is estimated to have a relative expanded uncertainty ranging at a 95 % confidence level from 1 % to 4 % depending on the temperature and pressure, with larger uncertainties in the critical region., Competing Interests: The authors declare no competing financial interest.

This paper presents new wide-ranging correlations for the viscosity and thermal conductivity of n -undecane based on critically evaluated experimental data. The correlations are designed to be used with a recently published equation of state that is valid from the triple point to 700 K, at pressures up to 500 MPa, with densities below 776.86 kg m -3 . The estimated uncertainty for the dilute-gas viscosity is 2.4%, and the estimated uncertainty for viscosity in the liquid phase for pressures up to 60 MPa over the temperature range 260 K to 520 K is 5%. The estimated uncertainty is 3% for the thermal conductivity of the low-density gas, and 3% for the liquid over the temperature range from 284 K to 677 K at pressures up to 400 MPa. Both correlations behave in a physically reasonable manner when extrapolated to the full range of the equation of state, however care should be taken when using the correlations outside of the validated range. The uncertainties will be larger outside of the validated range, and also in the critical region.

The available experimental data for the thermal conductivity of liquid bismuth, cobalt, germanium and silicon have been critically examined with the intention of establishing thermal conductivity reference correlations. All experimental data have been categorized into primary and secondary data according to the quality of measurement specified by a series of criteria. The proposed standard reference correlations for the thermal conductivity of liquid bismuth, cobalt, germanium, and silicon are respectively characterized by uncertainties of 10, 15, 16 and 9.5% at the 95% confidence level.

This paper presents new wide-ranging correlations for the viscosity and thermal conductivity of ethyl fluoride (R161) based on critically evaluated experimental data. The correlations are designed to be used with a recently published equation of state that is valid from 130 K to 450 K, at pressures up to 100 MPa. The estimated uncertainty at a 95% confidence level is 2% for the viscosity of low-density gas (pressures below 0.5 MPa), and 3% for the viscosity of the liquid over the temperature range from 243 K to 363 K at pressures up to 30 MPa. The estimated uncertainty is 3% for the thermal conductivity of the low-density gas, and 3% for the liquid over the temperature range from 234 K to 374 K at pressures up to 20 MPa. Both correlations may be used over the full range of the equation of state, but the uncertainties will be larger, especially in the critical region.

New, wide-range reference equations for the thermal conductivity of cyclohexane as a function of temperature and density are presented. The equations are based in part upon a body of experimental data that has been critically assessed for internal consistency and for agreement with theory whenever possible. We estimate the uncertainty (at the 95% confidence level) for the thermal conductivity of cyclohexane from the triple point (279.86 K) to 650 K at pressures up to 175 MPa to be 4% for the compressed liquid and supercritical phases. For the low-pressure gas phase (up to 0.1 MPa) over the temperature range 280 K to 680 K, the estimated uncertainty is 2.5%. Uncertainties in the critical region are much larger, since the thermal conductivity approaches infinity at the critical point and is very sensitive to small changes in density.

The available experimental data for the thermal conductivity of liquid copper, gallium, indium, iron, lead, nickel, and tin has been critically examined with the intention of establishing thermal conductivity reference correlations. All experimental data have been categorized into primary and secondary data according to the quality of measurement specified by a series of criteria. The proposed standard reference correlations for the thermal conductivity of liquid copper, gallium, indium, iron, lead, nickel, and tin are respectively characterized by uncertainties of 9.8, 15.9, 9.7, 13.7, 16.9, 7.7, and 12.6% at the 95% confidence level.

Due to concerns about global warming, there is interest in 2,3,3,3-Tetrafluoroprop-1-ene (R1234yf) and trans -1,3,3,3-Tetrafluoropropene (R1234ze(E)) as potential replacements for refrigerants with high global warming potential (GWP). In this manuscript we survey available data and provide viscosity correlations that cover the entire fluid range including vapor, liquid, and supercritical regions. The correlation for R1234yf is valid from the triple point (220 K) to 410 K at pressures up to 30 MPa, and the correlation for R1234ze(E) is valid from the triple point (169 K) to 420 K at pressures up to 100 MPa. The estimated uncertainty for both correlations at a 95 % confidence level is 2 % for the liquid phase over the temperature range 243 K to 363 K at pressures to 30 MPa, and 3 % for the gas phase at atmospheric pressure.

Objective: To compare the biomechanical properties of simulated humeral condylar fractures reduced with one of two screw fixation methods: 3.0 mm headless compression screw (HCS) or 3.5 mm cortical bone screw (CBS) placed in lag fashion., Methods: Bilateral humeri were collected from nine canine cadavers. Standardized osteotomies were stabilized with 3.0 mm HCS in one limb and 3.5 mm CBS in the contralateral limb. Condylar fragments were loaded to walk, trot, and failure loads while measuring construct properties and condylar fragment motion., Results: The 3.5 mm CBS-stabilized constructs were 36% stiffer than 3.0 mm HCS-stabilized constructs, but differences were not apparent in quality of fracture reduction nor in yield loads, which exceeded expected physiological loads during rehabilitation. Small residual fragment displacements were not different between CBS and HCS screws. Small fragment rotation was not significantly different between screws, but was weakly correlated with moment arm length (R² = 0.25)., Clinical Significance: A CBS screw placed in lag fashion provides stiffer fixation than an HCS screw, although both screws provide similar anatomical reduction and yield strength to condylar fracture fixation in adult canine humeri.

New, wide-range reference equations for the thermal conductivity of ethene and propene as a function of temperature and density are presented. The equations are based in part upon a body of experimental data that has been critically assessed for internal consistency and for agreement with theory whenever possible. For ethene, we estimate the uncertainty (at the 95% confidence level) for the thermal conductivity from 110 K to 520 K at pressures up to 200 MPa to be 5% for the compressed liquid and supercritical phases. For the low-pressure gas phase (to 0.1 MPa) over the temperature range 270 K to 680 K, the estimated uncertainty is 4%. The correlation is valid from 110 K to 680 K and up to 200 MPa, but it behaves in a physically reasonable manner down to the triple point and may be used at pressures up to 300 MPa, although the uncertainty will be larger in regions where experimental data were unavailable. In the case of propene, data are much more limited. We estimate the uncertainty for the thermal conductivity of propene from 180 K to 625 K at pressures up to 50 MPa to be 5% for the gas, liquid, and supercritical phases. The correlation is valid from 180 K to 625 K and up to 50 MPa, but it behaves in a physically reasonable manner down to the triple point and may be used at pressures up to 100 MPa, although the uncertainty will be larger in regions where experimental data were unavailable. For both fluids, uncertainties in the critical region are much larger, since the thermal conductivity approaches infinity at the critical point and is very sensitive to small changes in density.

This paper contains new, representative reference equations for the thermal conductivity of carbon dioxide. The equations are based in part upon a body of experimental data that has been critically assessed for internal consistency and for agreement with theory whenever possible. In the case of the dilute-gas thermal conductivity, we incorporated recent theoretical calculations to extend the temperature range of the experimental data. Moreover, in the critical region, the experimentally observed enhancement of the thermal conductivity is well represented by theoretically based equations containing just one adjustable parameter. The correlations are applicable for the temperature range from the triple point to 1100 K and pressures up to 200 MPa. The overall uncertainty (at the 95% confidence level) of the proposed correlation varies depending on the state point from a low of 1% at very low pressures below 0.1 MPa between 300 K and 700 K, to 5% at the higher pressures of the range of validity.

The primary objectives of this work were to formulate, blend, and characterize a set of four ultralow-sulfur diesel surrogate fuels in quantities sufficient to enable their study in single-cylinder-engine and combustion-vessel experiments. The surrogate fuels feature increasing levels of compositional accuracy (i.e., increasing exactness in matching hydrocarbon structural characteristics) relative to the single target diesel fuel upon which the surrogate fuels are based. This approach was taken to assist in determining the minimum level of surrogate-fuel compositional accuracy that is required to adequately emulate the performance characteristics of the target fuel under different combustion modes. For each of the four surrogate fuels, an approximately 30 L batch was blended, and a number of the physical and chemical properties were measured. This work documents the surrogate-fuel creation process and the results of the property measurements.

Objective: To investigate: 1) acanthocytosis and presence of acanthocytes in peritoneal fluid as a diagnostic marker for hemangiosarcoma (HSA) in dogs with non-traumatic hemoabdomen; and 2) the association between other erythrocyte, biochemical, and hematologic abnormalities as a mean of differentiating HSA from other disease., Study Design: Prospective double-blinded cohort study., Animals: Dogs (n = 40) with non-traumatic hemoabdomen., Methods: Dogs diagnosed with hemoabdomen (January 2012 to May 2013) had cytologic evaluation of abdominal effusion and peripheral blood smears. Peripheral blood CBC, PT, and aPTT, as well as blood and effusion acanthocytes, keratocytes, schistocytes, lactate, glucose, PCV, and TP results were compared using the paired t-test or Fisher's exact test. Based on histologic confirmation of HSA, dogs were divided into 2 groups (HSA, non-HSA) and variables compared., Results: There was no significant difference in erythrocyte morphology in abdominal effusion or peripheral blood between dogs with HSA or non-HSA related hemoabdomen. Platelet concentration and peripheral blood PCV were significantly lower in the HSA group., Conclusions: A reliable preoperative biochemical or cytologic test to differentiate between HSA and non-HSA related hemoabdomen was not identified., (© Copyright 2015 by The American College of Veterinary Surgeons.)

Sample preparation prior to analysis by liquid chromatography electrospray ionization mass spectrometry (LC-ESI-MS) usually involves the storage of frozen peptide samples in an acidic environment for variable time periods. Questions arose in our laboratory regarding the stability of peptides in acid under medium- to long-term storage. Thus, a 10-month longitudinal study was designed to assess the effect on storage of tryptic peptides at -20 and -80°C under acidic conditions. Our conclusion and proposal from this evaluation is that the optimal storage conditions of peptide samples in acid for proteomic experiments is at -80°C and, ideally, as separate aliquots., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Affinity purification coupled to 1-D gel-free liquid chromatography mass spectrometry (LC-MS) is a well-established and widespread approach for the analyses of noncovalently interacting protein complexes. In this study, two proteins conjugated to a streptavidin-binding peptide and hemagglutinin double tag were expressed in the respective Flp-In HEK293 cell lines: green fluorescent protein (SH-GFP) and TANK binding kinase 1 (SH-TBK1&#95;MOUSE). Fluorescent anti-HA immunoblots revealed that the expression level of SH-GFP was ∼50% lower than that of SH-TBK1&#95;MOUSE. Subsequently, the input material was normalized to obtain a similar quantity of purified SH-tagged proteins. Optimization of the release of protein complexes from the anti-HA-agarose with different eluting agents was then assessed. With respect to the total number of protein groups identified in the purified complexes, elution with 2% SDS surpassed both 100 mM glycine and 100 mM formic acid. Relative quantitation of the purified protein complexes using TMT 6-plex reagents confirmed the higher efficiency of the 2% SDS elution followed by filter-aided sample preparation (FASP). The data presented in this study provide a new application of FASP to quantitative MS analysis of affinity-purified protein complexes. We have termed the approach abFASP-MS, or affinity-based filter-aided sample preparation mass spectrometry.

The identification and validation of cross-linked peptides by mass spectrometry remains a daunting challenge for protein-protein cross-linking approaches when investigating protein interactions. This includes the fragmentation of cross-linked peptides in the mass spectrometer per se and following database searching, the matching of the molecular masses of the fragment ions to the correct cross-linked peptides. The hybrid linear trap quadrupole (LTQ) Orbitrap Velos combines the speed of the tandem mass spectrometry (MS/MS) duty circle with high mass accuracy, and these features were utilized in the current study to substantially improve the confidence in the identification of cross-linked peptides. An MS/MS method termed multiple and sequential data acquisition method (MSDAM) was developed. Preliminary optimization of the MS/MS settings was performed with a synthetic peptide (TP1) cross-linked with bis[sulfosuccinimidyl] suberate (BS(3)). On the basis of these results, MSDAM was created and assessed on the BS(3)-cross-linked bovine serum albumin (BSA) homodimer. MSDAM applies a series of multiple sequential fragmentation events with a range of different normalized collision energies (NCE) to the same precursor ion. The combination of a series of NCE enabled a considerable improvement in the quality of the fragmentation spectra for cross-linked peptides, and ultimately aided in the identification of the sequences of the cross-linked peptides. Concurrently, MSDAM provides confirmatory evidence from the formation of reporter ions fragments, which reduces the false positive rate of incorrectly assigned cross-linked peptides.

The performance of two proteomic sample preparation methods, "pseudoshotgun" (PSG) and filter-aided sample preparation (FASP) were compared in terms of the number of identified proteins, representation of cellular component GO (gene ontology) categories in the obtained list of proteins, and the efficiency of both methods in the proteomic analysis of a very low number of cells. Both methods were combined to obtain a proteomic profile of a short-term culture (passage 3) of melanoma cells, established in our laboratory from a human metastatic melanoma lesion. The data revealed that with FASP, usually more proteins are identified than with PSG when analyzing a higher number of cells (≥ 5000/injection), whereas PSG is favorable when analyzing only a very small amount of cells (250-500/injection). PSG and FASP, however, are complementary techniques, as combining both methods further increases the number of identified proteins. Moreover, we show that it is feasible to identify a substantial number of proteins from only 250 cells/injection that is equivalent to 60 ng of protein.

Protein complexes form, dissociate and re-form in order to perform specific cellular functions. In this two-pronged protocol, noncovalent protein complexes are initially isolated by affinity purification for subsequent identification of the components by liquid chromatography high-resolution mass spectrometry (LC-MS) on a hybrid LTQ Orbitrap Velos. In the second prong of the approach, the affinity-purification strategy includes a chemical cross-linking step to 'freeze' a series of concurrently formed, heterogeneous protein subcomplex species that are visualized by gel electrophoresis. This branch of the methodology amalgamates standard and well-practiced laboratory methods to reveal compositional changes that occur in protein complex architecture. By using mouse N-terminally tagged streptavidin-binding peptide-hemagglutinin-TANK-binding kinase 1 (SH-TBK1), we chemically cross-linked the affinity-purified complex of SH-TBK1 with the homobifunctional lysine-specific reagent bis(sulfosuccinimidyl) suberate (BS(3)), and we separated the resultant protein complexes by denaturation and by silver-stained one- and two-dimensional SDS-PAGE. We observed a range of cross-linked TBK1 complexes of variable pI and M(r) and confirmed them by immunoblotting. LC-MS analysis of in situ-digested cross-linked proteins shows differences in the composition of the TBK1 subcomplexes. The protocol is inherently simple and can be readily extended to the investigation of a range of protein complexes. From cell lysis to data generation by LC-MS, the protocol takes approximately 2.5 to 5.5 d to perform.

The 5-hydroxytryptamine 1A receptor (serotonin 1A receptor; 5-HT(1A) R) is involved in a large series of brain functions, and roles in anxiety, depression, and cognition have been reported. So far, published information on mass spectrometrical characterization of 5-HT(1A) R is limited to the presence of two 5-HT(1A) R peptides in rat's whole brain as observed by in-solution digestion followed by LC-MS/MS. Knowledge about the protein sequence and PTMs, however, would have implications for generation of specific antibodies and designing studies on the 5-HT(1A) R at the protein level. A rat recombinant 5-HT(1A) R was extracted from the tsA201 cell line, run using several gel-based principles with subsequent in-gel digestion with several proteases, chymotrypsin, trypsin, AspN, proteinase K, and pepsin followed by nano-LC-ESI-MS/MS analysis on a high capacity ion trap and an LTQ Orbitrap Velos. Using two search engines, Mascot and Modiro™, the recombinant 5-HT(1A) R was identified showing 94.55% sequence coverage. A single phosphorylation at S301 was identified and verified by phosphatase treatment and a series of amino acid substitutions were detected. Characterization of 5-HT(1A) R, a key player of brain functions and neurotransmission, was shown and may enable generation of specific antibodies, design of future, and interpretation of previous studies in the rat at the protein level., (© 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Sipuleucel-T was approved by the US Food and Drug Administration on April 29, 2010, as an immunotherapy for late-stage prostate cancer. To manufacture sipuleucel-T, mononuclear cells harvested from the patient are incubated with a recombinant prostatic acid phosphatase (PAP) antigen and reinfused. The manufacturer proposes that antigen-presenting cells exogenously activated by PAP induce endogenous T-cells to attack PAP-bearing prostate cancer cells. However, the lack of demonstrable tumor responses has prompted calls for scrutiny of the design of the trials in which sipuleucel-T demonstrated a 4-month survival benefit. Previously unpublished data from the sipuleucel-T trials show worse overall survival in older vs younger patients in the placebo groups, which have not been shown previously to be prognostic for survival in castration-resistant prostate cancer patients receiving chemotherapy. Because two-thirds of the cells harvested from placebo patients, but not from the sipuleucel-T arm, were frozen and not reinfused, a detrimental effect of this large repeated cell loss provides a potential alternative explanation for the survival "benefit." Patient safety depends on adequately addressing this alternative explanation for the trial results.

The recent National Academy of Sciences report on forensic sciences states that the study of fire patterns and debris in arson fires is in need of additional work and eventual standardization. We discuss a recently introduced method that can provide predicted evaporation patterns for ignitable liquids as a function of temperature. The method is a complex fluid analysis protocol, the advanced distillation curve approach, featuring a composition explicit data channel for each distillate fraction (for qualitative, quantitative, and trace analysis), low uncertainty temperature measurements that are thermodynamic state points that can be modeled with an equation of state, consistency with a century of historical data, and an assessment of the energy content of each distillate fraction. We discuss the application of the method to kerosenes and gasolines and outline how expansion of the scope of fluids to other ignitable liquids can benefit the criminalist in the analysis of fire debris for arson., (© 2010 American Academy of Forensic Sciences.)

Objective: To compare the bending structural stiffness and bending strength of thick and thin 2.4 mm limited contact dynamic compression plates (2.4 LC-DCP), 2.0 mm LC-DCP (2.0 LC-DCP), and 2.0 dynamic compression plates (2.0 DCP)., Study Design: In vitro mechanical study., Methods: Two thicknesses of 2.4 LC-DCP, 2.0 LC-DCP, and 2.0 DCP stainless-steel plates were tested in 4-point bending. Data were collected during bending until implants plastically deformed. Bending structural stiffness and bending strength were determined from load displacement curves. Mechanical properties were compared between plates and the effects of plate type, size, and thickness on stiffness and strength were assessed using ANOVA., Results: The thick 2.4 LC-DCP implant was the stiffest and strongest; the thin 2.0 DCP implant was most compliant and weakest. Larger sized plates, thicker plates, and limited contact design of plates enhanced stiffness and strength. For the plates studied, plate size had a larger effect than plate type or thickness on stiffness and strength., Conclusion: Increasing the size (width) and thickness of plates increases both the bending structural stiffness and strength. For the plates studied, LC-DCP implants were stiffer and stronger than DCP implants., Clinical Relevance: Plate bending structural stiffness and strength can be most effectively enhanced by using a larger sized plate, but gains can also be achieved by using a thicker plate and/or an LC-DCP instead of a DCP implant when possible., (© Copyright 2010 by The American College of Veterinary Surgeons.)

The analysis of complex fluids such as crude oils, fuels, vegetable oils and mixed waste streams poses significant challenges arising primarily from the multiplicity of components, the different properties of the components (polarity, polarizability, etc.) and matrix properties. We have recently introduced an analytical strategy that simplifies many of these analyses, and provides the added potential of linking compositional information with physical property information. This aspect can be used to facilitate equation of state development for the complex fluids. In addition to chemical characterization, the approach provides the ability to calculate thermodynamic properties for such complex heterogeneous streams. The technique is based on the advanced distillation curve (ADC) metrology, which separates a complex fluid by distillation into fractions that are sampled, and for which thermodynamically consistent temperatures are measured at atmospheric pressure. The collected sample fractions can be analyzed by any method that is appropriate. The analytical methods we have applied include gas chromatography (with flame ionization, mass spectrometric and sulfur chemiluminescence detection), thin layer chromatography, FTIR, corrosivity analysis, neutron activation analysis and cold neutron prompt gamma activation analysis. By far, the most widely used analytical technique we have used with the ADC is gas chromatography. This has enabled us to study finished fuels (gasoline, diesel fuels, aviation fuels, rocket propellants), crude oils (including a crude oil made from swine manure) and waste oils streams (used automotive and transformer oils). In this special issue of the Journal of Chromatography, specifically dedicated to extraction technologies, we describe the essential features of the advanced distillation curve metrology as an analytical strategy for complex fluids., (Published by Elsevier B.V.)

An equation for the density of hydrogen gas has been developed that agrees with the current standard to within 0.01 % from 220 K to 1000 K with pressures up to 70 MPa, to within 0.01 % from 255 K to 1000 K with pressures to 120 MPa, and to within 0.1 % from 200 K to 1000 K up to 200 MPa. The equation is a truncated virial-type equation based on pressure and temperature dependent terms. The density uncertainty for this equation is the same as the current standard and is estimated to be 0.04 % (combined uncertainty with a coverage factor of 2) between 250 K and 450 K for all pressures, and 0.1 % for lower temperatures. Comparisons are presented with experimental data and with the full equation of state.

We study the uniformly weighted ensemble of force balanced configurations on a triangular network of nontensile contact forces. For periodic boundary conditions corresponding to isotropic compressive stress, we find that the probability distribution for single-contact forces decays faster than exponentially. This superexponential decay persists in lattices diluted to the rigidity percolation threshold. On the other hand, for anisotropic imposed stresses, a broader tail emerges in the force distribution, becoming a pure exponential in the limit of infinite lattice size and infinitely strong anisotropy.

Objective: To assess the accuracy of current antemortem and postmortem techniques for determining tracheal luminal stenosis., Animals: 15 dogs., Procedure: Percentage of tracheal luminal stenosis (PTLS) was determined by 6 methods, using measurements obtained by radiography, tracheoscopy, and necropsy after selected tracheostomy techniques were performed. To calculate PTLS, dorsoventral tracheal diameter was measured from preoperative and postoperative lateral cervical radiographic views. Preoperative or normal tracheal segments adjacent to the stenotic area were used to obtain normal tracheal diameter measurements. Planimetrically determined cross-sectional area (CSA), obtained from pre- and postoperative tracheoscopic photographs, was used to calculate PTLS. The CSA of tracheal specimens obtained at necropsy was determined, using the formula for an ellipse. Percentage of luminal stenosis was calculated, using CSA of the stenotic site and of segments craniad and caudad to the site obtained at necropsy or at surgery. All methods were compared with the control method of planimetrically determined CSA of sections obtained at necropsy of the tracheostomy and segments craniad and caudad to the site., Results: Correlation was poor for radiographic and tracheoscopic techniques (r = 0.146 to 0.458, P > 0.05) The formula for an ellipse accurately predicted PTLS when measurements obtained at surgery (r = 0.516, P = 0.049) or segments craniad and caudad (r = 0.853, P < 0.001) to the site were used., Conclusion: Antemortem methods of assessing PTLS did not correlate with control planimetric methods. Methods using CSA determined by tracheal diameter were weakly correlated to control planimetric techniques., Clinical Relevance: Accurate measurement of the degree of tracheal stenosis cannot be made in clinical patients using current techniques.

Three branched-chain fatty acids (7-hydroxy-4,6-dimethylnona-2,4-dienoic acid [compound 1], its 7-epimer [compound 2], and 7-keto-4,6-dimethylnona-2,4-dienoic acid [compound 3]) and a ketone (9-hydroxy-6,8-dimethylundeca-4,6-dien-3-one [compound 4]) were isolated from the culture broth of mutants of Streptomyces fradiae which were blocked in the biosynthesis of the macrolide antibiotic tylosin. Two phenotypic classes of mutants of this organism which were blocked in the addition of mycaminose to tylactone (compound 6) accumulated these compounds. These compounds were not produced by mutants which were blocked in lactone synthesis, in steps beyond mycaminose addition, or by the wild-type strain. Synthesis of these compounds, like synthesis of tylosin, was inhibited by the addition of cerulenin. Compounds 1, 2, and 3 were partially interconvertible by these mutants; but they were not produced from the degradation of tylactone and they were not directly incorporated into tylosin by intact cells. The structures of compounds 1 and 2 were equivalent to that of a predicted intermediate (S. Yue, J. S. Duncan, Y. Yamamoto, and C. R. Hutchinson, J. Am. Chem. Soc. 109:1253-1255, 1987) in the biosynthesis of tylactone. The ketone (compound 4) reported previously (N. D. Jones, M. O. Chaney, H. A. Kirst, G. M. Wild, R. H. Baltz, R. L. Hamill, and J. W. Paschal, J. Antibiot. 35:420-425, 1982) appears to be the decarboxylation product of the intermediate following that represented by compound 1. This represents the first report of the isolation of putative precursors of tylactone from tylosin-producing organisms.

Several cosmid clones from Streptomyces ambofaciens containing the spiramycin resistance gene srmB were introduced into S. fradiae PM73, a mutant defective in tylosin synthesis, resulting in tylosin synthesis. The DNA responsible for this complementation was localized to a 10.5-kilobase EcoRI fragment. A 32-kilobase DNA segment which included the srmB spiramycin resistance gene and DNA which complemented the defect in strain PM73 were mutagenized in vivo with Tn10 carrying the gene for Nmr (which is expressed in Streptomyces spp.) or in vitro by insertional mutagenesis with a drug resistance gene (Nmr) cassette. When these mutagenized DNA segments were crossed into the S. ambofaciens chromosome, three mutant classes blocked in spiramycin synthesis were obtained. One mutant accumulated two precursors of spiramycin, platenolide I and platenolide II. Two mutants, when cofermented with the platenolide-accumulating mutant, produced spiramycin. Tylactone supplementation of these two mutants resulted in the synthesis of a group of compounds exhibiting antibiotic activity. Two other mutants failed to coferment with any of the other mutants or to respond to tylactone supplementation.