Δ9-tetrahydrocannabinol (THC), the main psychoactive compound in cannabis, and other drug molecules that have large molar masses, are often described as 'nonvolatile' and are presumed to be carried in exhaled breath aerosols. Large variabilities in THC concentrations in breath have been measured with devices that only collect aerosols; it is possible that neglecting the vapor phase could be responsible. Partitioning of compounds between vapor and aerosol phases is directly dependent on vapor pressure (psat), which itself is strongly dependent on temperature. We describepsatmeasurements for THC, cannabidiol (CBD), and cannabinol (CBN) using a gas-saturation apparatus. The measured values ofpsatfor 364 K to 424 K are 0.0459 Pa to 7.833 Pa for THC, 0.0826 Pa to 13.44 Pa for CBD, and 0.0199 Pa to 5.678 Pa for CBN. The combined standard (k= 1, 68% confidence) measurement uncertainty inpsatranges from 2.9% to 5.3% for CBD and CBN, and from 5.2% to 9.5% for THC. To obtain thepsatat human body and exhaled breath temperatures, we extrapolated the measurements for each cannabinoid with a thermodynamic correlation. Then a vapor-aerosol partitioning model was used to predict mole fractions of each cannabinoid in each phase of exhaled breath. All three cannabinoids were predicted to reside primarily in the vapor phase of exhaled breath. However, relatively small changes in temperature or aerosol concentration can significantly impact the predicted partitioning. This work illustrates the utility of low-uncertaintypsatmeasurements for any drug, including those thought to be too low in volatility for vapor-phase sampling, and may extend the market for forensic drug tests and clinical diagnostic tests via breath analysis.

Halogenated non-innocent vanadium( v ) Schiff base mono-substituted catecholates have a low stability and anti-proliferative properties similar to vanadate. A linear correlation between complex redox potential and the catechol p K a was observed.

Cannabinoids can be detected in breath after cannabis use, but different breath matrices need to be explored as studies to date with filter-based devices that collect breath aerosols have not demonstrated that breath-based measurements can reliably identify recent cannabis use. Exhaled breath condensate (EBC) is an unexplored aqueous breath matrix that contains condensed volatile compounds and water vapor in addition to aerosols. EBC was collected from participants both before and at two time points (0.7 ± 0.2 h and 1.7 ± 0.3 h) after observed cannabis use. Eleven different cannabinoids were monitored with liquid chromatography tandem mass spectrometry. Five different cannabinoids, including Δ9-tetrahydrocannabinol (THC), were detected in EBC collected from cannabis users. THC was detected in some EBC samples before cannabis use, despite the requested abstinence period. THC was detected in all EBC samples collected at 0.7 h post use and decreased for all participants at 1.7 h. Non-THC cannabinoids were only detected after cannabis use. THC concentrations in EBC samples collected at 0.7 h showed no trend with sample metrics like mass or number of breaths. EBC sampling devices deserve further investigation with respect to modes of cannabis use (e.g, edibles), post use time points, and optimization of cannabinoid recovery.

Vapor pressure (psat) 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 psat measurements, known as dynamic vapor microextraction (DVME), was used to measure psat 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 psat 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·psat to 0.0584·psat, 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 psat that are 200 times higher than in the DVME validation study with n-eicosane (C20H42). The oxidative stability of linalool was improved by the addition of 0.2 mass % of the antioxidant tert-butylhydroquinone.

An accurate cannabis breathalyzer based on quantitation of the psychoactive cannabinoid Δ 9 -tetrahydrocannabinol (THC) could be an important tool for deterring impaired driving. Such a device does not exist. Simply translating what is known about alcohol breathalyzers is insufficient because ethanol is detected as a vapor. THC has extremely low volatility and is hypothesized to be carried in breath by aerosol particles formed from lung surfactant. Exhaled breath aerosols can be recovered from electrostatic filter devices, but consistent quantitative results across multiple studies have not been demonstrated. We used a simple-to-use impaction filter device to collect breath aerosols from participants before and after they smoked a legal market cannabis flower containing ∼25% Δ 9 -tetrahydrocannabinolic acid. Breath collection occurred at an intake session (baseline-intake) and four weeks later in a federally-compliant mobile laboratory 15 min before (baseline-experimental) and 1 h after cannabis use (post-use). Cannabis use was in the participant’s residence. Participants were asked to follow a breathing maneuver designed to increase aerosol production. Breath extracts were analyzed by liquid chromatography with tandem mass spectrometry with multiple reaction monitoring of two transitions for analytes and their deuterated internal standards. Over more than 1 yr, 42 breath samples from 18 participants were collected and analyzed in six batches. THC was quantified in 31% of baseline-intake, 36% of baseline-experimental, and 80% of 1 h post-use breath extracts. The quantities observed 1 h post-use are compared to those reported in six other pilot studies that sampled breath at known intervals following cannabis use and are discussed with respect to participant characteristics and breath sampling protocols. Larger studies with verified abstinence and more post-use timepoints are necessary to generate statistically significant data to develop meaningful cannabis breathalyzer technology.

A hydrophobic Schiff base catecholate vanadium complex was recently discovered to have anticancer properties superior to cisplatin and suited for intratumoral administration. This [VO(HSHED)(DTB)] complex, where HSHED is N-(salicylideneaminato)-N'-(2-hydroxyethyl)-1,2-ethanediamine and the non-innocent catecholato ligand is di-t-butylcatecholato (DTB), has higher stability compared to simpler catecholato complexes. Three new chloro-substituted Schiff base complexes of vanadium(V) with substituted catecholates as co-ligands were synthesized for comparison with their non-chlorinated Schiff base vanadium complexes, and their properties were characterized. Up to four geometric isomers for each complex were identified in organic solvents using 51V and 1H NMR spectroscopies. Spectroscopy was used to characterize the structure of the major isomer in solution and to demonstrate that the observed isomers are exchanged in solution. All three chloro-substituted Schiff base vanadium(V) complexes with substituted catecholates were also characterized by UV-vis spectroscopy, mass spectrometry, and electrochemistry. Upon testing in human glioblastoma multiforme (T98g) cells as an in vitro model of brain gliomas, the most sterically hindered, hydrophobic, and stable compound [t1/2 (298 K) = 15 min in cell medium] was better than the two other complexes (IC50 = 4.1 ± 0.5 μM DTB, 34 ± 7 μM 3-MeCat, and 19 ± 2 μM Cat). Furthermore, upon aging, the complexes formed less toxic decomposition products (IC50 = 9 ± 1 μM DTB, 18 ± 3 μM 3-MeCat, and 8.1 ± 0.6 μM Cat). The vanadium complexes with the chloro-substituted Schiff base were more hydrophobic, more hydrolytically stable, more easily reduced compared to their corresponding parent counterparts, and the most sterically hindered complex of this series is only the second non-innocent vanadium Schiff base complex with a potent in vitro anticancer activity that is an order of magnitude more potent than cisplatin under the same conditions.

The interpeptidic CuII exchange rate constants were measured for two Cu amyloid-β complexes, Cu(Aβ1-16) and Cu(Aβ1-28), to fluorescent peptides GHW and DAHW using a quantitative tryptophan fluorescence quenching methodology. The second-order rate constants were determined at three pH values (6.8, 7.4, and 8.7) important to the two Cu(Aβ) coordination complexes, components Cu(Aβ)I and Cu(Aβ)II. The interpeptidic CuII exchange rate constant is approximately 104 M-1 s-1 but varies in magnitude depending on many variables. These include pH, length of the Aβ peptide, location of the anchoring histidine ligand in the fluorescent peptide, number of amide deprotonations required in the tryptophan peptide to coordinate CuII, and interconversion between Cu(Aβ)I and Cu(Aβ)II. We also present EPR data probing the CuII exchange between peptides and the formation of ternary species between Cu(Aβ) and GHW. As the nonfluorescent GHK and DAHK peptides are important motifs found in the blood and serum, their ability to sequester CuII ions from Cu(Aβ) complexes may be relevant for the metal homeostasis and its implication in Alzheimer's disease. Thus, their kinetic CuII interpeptidic exchange rate constants are important chemical rate constants that can help elucidate the complex CuII trafficking puzzle in the synaptic cleft.

We sought to determine if 28 days of probiotic supplementation influenced the plasma amino acid (AA) response to acute whey protein feeding. METHODS: Twenty-two recreationally active men (n = 11; 24.3 ± 3.2 yrs; 89.3 ± 7.2 kg) and women (n = 11; 23.0 ± 2.8 yrs; 70.2 ± 15.2 kg) participated in this double-blind, placebo-controlled, randomized study. Before (PRE) and after 28 days of supplementation (POST), participants reported to the lab following a 10-hr fast and provided a resting blood draw (0 min), then subsequently consumed 25 g of whey protein. Blood samples were collected at 15-min intervals for 2 h post-consumption (15–120 min) and later analyzed for plasma leucine, branched-chain AA (BCAA), essential AA (EAA), and total AA (TAA). Participants received a probiotic (PROB) consisting of 1 x10-9 colony forming units (CFU) Bacillus subtilis DE111 (n = 11) or a maltodextrin placebo (PL) (n = 11) for 28 days. Plasma AA response and area under the curve (AUC) values were analyzed via repeated measures analysis of variance. RESULTS: Our analysis indicated no significant (p < 0.05) differential responses for plasma leucine, BCAA, EAA, or TAA between PROB and PL from PRE to POST. AUC analysis revealed no group × time interaction for plasma leucine (p = 0.524), BCAA (p = 0.345), EAA (p = 0.512), and TAA (p = 0.712). CONCLUSION: These data indicate that 28 days of Bacillus subtilis DE111 does not affect plasma AA appearance following acute whey protein ingestion.

Dynamic vapor microextraction (DVME) is a new method that enables rapid vapor pressure measurements on large molecules with state-of-the-art measurement uncertainty for vapor pressures near 1 Pa. Four key features of DVME that allow for the rapid collection of vapor samples under thermodynamic conditions are (1) the use of a miniature vapor-equilibration vessel (the "saturator") to minimize the temperature gradients and internal volume, (2) the use of a capillary vapor trap to minimize the internal volume, (3) the use of helium carrier gas to minimize nonideal mixture behavior, and (4) the direct measurement of pressure inside the saturator to accurately account for overpressure caused by viscous flow. The performance of DVME was validated with vapor pressure measurements of n-eicosane (C20H42) at temperatures from 344 to 374 K. A thorough uncertainty analysis indicated a relative standard uncertainty of 2.03-2.82% for measurements in this temperature range. The measurements were compared to a reference correlation for the vapor pressures of n-alkanes; the deviation of the measurements from the correlation was ≤2.85%. The enthalpy of vaporization of n-eicosane at 359.0 K was calculated to be ΔvapH = 91.27 ± 0.28 kJ/mol compared to ΔvapH(corr) = 91.44 kJ/mol for the reference correlation. Total measurement periods as short as 15 min (3 min of thermal equilibration plus 12 min of carrier gas flow) were shown to be sufficient for high-quality vapor pressure measurements at 364 K.

PURPOSE: The aim of the current study was to examine the efficacy of whey protein ingestion with or without a protease enzyme complex on amino acid (AA) availability following acute lower-body resistance exercise. METHODS: Ten resistance trained men (24.4±4.1yr, 179.1±8.6cm, 92.6±10.4kg) with at least one year of resistance training experience volunteered to participate in this placebo-controlled, randomized, cross-over designed study. Following an overnight fast, participants performed lower-body acute resistance exercises consisting of four sets each of the leg press and leg extension exercises (8-10 repetitions at 75% of 1RM) followed by consumption of one of three drinks of equivalent volume, taste, and appearance which consisted of either: (a) 26g whey protein + 250 mg protease supplement + whey (PW) (b) 26g whey protein (W), or (c) a non-caloric flavored water drink (PL). Blood samples were collected before exercise, immediately-post (0min), 30-, 60-, 90-, 120-, and 180-minutes post-exercise (30min, 60min, 90min, 120min, 180min respectively). Plasma amino acid samples were analyzed for essential (EAA), branched-chained (BCAA), and leucine concentrations via liquid chromatography-mass spectrometry (LC-MS). A 2-way repeated measures analysis of variance (ANOVA) was used to identify differences between treatments over time. Area under the curve was calculated via the trapezoidal technique and analyzed via a one-way ANOVA. RESULTS: Significant main effects for time (p<0.001) and time x group interactions (p<0.001) were found for leucine, BCAA, and EAA. PW drink resulted in significantly greater plasma leucine, BCAA and EAA concentrations at 30 min compared to PL (p<0.001) while not different than W. Leucine was significantly elevated at 30min (p=0.007) and EAAs at 180min (p=0.004) compared to 0min for PW. The AUC for PW and W were both significantly elevated for leucine, BCAAs and EAAs compared to PL (p<0.001). CONCLUSION: While no significant differences were found between the W and PW supplementation groups during the 3-hr period after resistance training; the PW group produced significantly greater leucine concentrations at 30min and for EAAs at 180min than PL compared to 0min. Results indicate that PW may provide a modest improvement of AA appearance in blood following acute resistance exercise.

This double-blind study examined effects of a protease enzyme blend (Prohydrolase®) added to whey protein on post-resistance exercise aminoacidemia and intramuscular anabolic signaling were investigated in ten resistance-trained males. Participants completed 4 sets of 8-10 repetitions in the leg press and leg extension exercises at 75% of 1-repetition maximum. Participants then consumed either 250 mg of Prohydrolase® + 26 g of whey protein (PW), 26 g whey alone (W), or non-nutritive control (CON) in counterbalanced order. Blood samples were obtained prior to exercise (baseline) and then immediately-post (IP), 30-, 60-, 90-, 120-, and 180-min post-exercise. Muscle biopsies were taken at baseline, 1-h (1H), and 3-h (3H) post-exercise. Phosphorylation of AKTSer437 was decreased (3H only: p < 0.001), mTORSer2448 was increased (1H: p = 0.025; 3H: p = 0.009), and p70S6KThr412 remained unchanged similarly for each condition. Plasma leucine, branch-chained amino acids, and essential amino acid concentrations for PW were significantly higher than CON (p < 0.05) at 30 min and similar to W. Compared to IP, PW was the only treatment with elevated plasma leucine levels at 30 min (p = 0.007; ∆ = 57.8 mmol/L, 95% Confidence Interval (CI): 20.0, 95.6) and EAA levels at 180 min (p = 0.003; ∆ = 179.1 mmol/L, 95% CI: 77.5, 280.7). Area under the curve amino acid analysis revealed no differences between PW and W. While no different than W, these data indicate that PW was the only group to produce elevated amino acid concentrations 30-min and 180-min post-ingestion.

Menaquinones (MKs) are essential for electron transport in prokaryotes, and importantly, partially saturated MKs represent a novel virulence factor. However, little is known regarding how the degree of saturation in the isoprenyl side chain influences conformation or quinone redox potential. MenJ is an enzyme that selectively reduces the second isoprene unit on MK-9 and is contextually essential for the survival of Mycobacterium tuberculosis in J774A.1 macrophage-like cells, suggesting that MenJ may be a conditional drug target for pathogenic mycobacteria. Therefore, fundamental information about the properties of this system is important, and we synthesized the simplest MKs, unsaturated MK-1 and the saturated analogue, MK-1(H2). Using two-dimensional nuclear magnetic resonance spectroscopy, we established that MK-1 and MK-1(H2) adopted similar folded-extended conformations (i.e., the isoprenyl side chain folds upward) in each solvent examined but the folded-extended conformations differed slightly between organic solvents. Saturation of the isoprenyl side chain slightly altered the MK-1 analogue conformation in each solvent. We used molecular mechanics to illustrate the MK-1 analogue conformations. The measured quinone redox potentials of MK-1 and MK-1(H2) differed between organic solvents (presumably due to differences in dielectric constants), and remarkably, an ∼20 mV semiquinone redox potential difference was observed between MK-1 and MK-1(H2) in pyridine, acetonitrile, and dimethyl sulfoxide, demonstrating that the degree of saturation in the isoprenyl side chain of MK-1 influences the quinone redox potential. Finally, MK-1 and MK-1(H2) interacted with Langmuir phospholipid monolayers and Aerosol-OT reverse micelle (RM) model membrane interfaces, where MK-1 adopted a slightly different folded conformation within the RM model membrane interface.

The light-induced photolysis of [Zn(NTAdeCage)]- generates a temporally controlled burst of Zn2+, which is rapidly chelated in situ by the free ligand Zincon2-. The [Zn(Zincon)]2- coordination progress is monitored using absorption spectroscopy in bulk aqueous buffer and reverse micelle environments. The [Zn(NTAdeCage)]- photocage and free ligand Zincon2- have different reverse micelle locations that affect the [Zn(Zincon)]2- formation at the nanoscale compared to the bulk aqueous buffer. The formation of [Zn(Zincon)]2- in a bulk aqueous buffer is more efficient despite the released Zn2+ and Zincon2- being physically closer within reverse micelles. The observed reduction of complex formation is attributed to the interfacial partitioning of Zincon2-, distinct from the Zn2+ photocage in the water pool, requiring diffusion for the species to meet to form [Zn(Zincon)]2-. This work introduces a proof-of-concept methodology to experimentally measure fast chelation reactions in confined spaces and thus provides an approach to exploring cellular responses.

The interpeptidic exchange of Cu(II) between biologically relevant peptides like Gly-His-Lys (GHK) was measured through proximity static fluorescence quenching of a noncoordinating tryptophan residue by Cu(II). The inability to spectrally distinguish between starting and final Cu(H-1GHK)+ complexes by the current methods was solved by the replacement of noncoordinating lysine for tryptophan in the starting complex, Cu(H-1GHW). Because the apoGHW is the only fluorescent species, the recovered fluorescence is directly proportional to the [Cu(II)]exchanged between GHW and GHK. The apparent second-order rate constants of the exchanges from Cu(H-1GHW) to GHK and DAHK are 1.6 (±0.2) × 102 and 5.0 (±0.7) × 101 M-1 s-1, respectively. The easy-to-implement kinetic fluorescent method described here for Cu(II) interpeptidic exchange can be expanded to other biological systems.

Menaquinones (naphthoquinones, MK) are isoprenoids that play key roles in the respiratory electron transport system of some prokaryotes by shuttling electrons between membrane-bound protein complexes acting as electron acceptors and donors. Menaquinone-2 (MK-2), a truncated MK, was synthesized, and the studies presented herein characterize the conformational and chemical properties of the hydrophobic MK-2 molecule. Using 2D NMR spectroscopy, we established for the first time that MK-2 has a folded conformation defined by the isoprenyl side-chain folding back over the napthoquinone in a U-shape, which depends on the specific environmental conditions found in different solvents. We used molecular mechanics to illustrate conformations found by the NMR experiments. The measured redox potentials of MK-2 differed in three organic solvents, where MK-2 was most easily reduced in DMSO, which may suggest a combination of solvent effect (presumably in part because of differences in dielectric constants) and/or conformational differences of MK-2 in different organic solvents. Furthermore, MK-2 was found to associate with the interface of model membranes represented by Langmuir phospholipid monolayers and Aerosol-OT (AOT) reverse micelles. MK-2 adopts a slightly different U-shaped conformation within reverse micelles compared to within solution, which is in sharp contrast to the extended conformations illustrated in literature for MKs.