Hyperpolarized lithium-6 as a sensor of nanomolar contrast agents.

Lithium is widely used in psychotherapy. The (6)Li isotope has a long intrinsic longitudinal relaxation time T(1) on the order of minutes, making it an ideal candidate for hyperpolarization experiments. In the present study we demonstrated that lithium-6 can be readily hyperpolarized within 30 min, while retaining a long polarization decay time on the order of a minute. We used the intrinsically long relaxation time for the detection of 500 nM contrast agent in vitro. Hyperpolarized lithium-6 was administered to the rat and its signal retained a decay time on the order of 70 sec in vivo. Localization experiments imply that the lithium signal originated from within the brain and that it was detectable up to 5 min after administration. We conclude that the detection of submicromolar contrast agents using hyperpolarized NMR nuclei such as (6)Li may provide a novel avenue for molecular imaging.

In vivo 1H NMR measurement of glycine in rat brain at 9.4 T at short echo time.

Glycine is an amino acid present in mammalian brain, where it acts as an inhibitory and excitatory neurotransmitter. The two detectable protons of glycine give rise to a singlet at 3.55 ppm that overlaps with the more intense myo-inositol resonances, and its measurement has traditionally required specific editing efforts. The aim of the current study was to reduce the signal intensity of myo-inositol relative to that of glycine by exploiting the fast signal J-evolution of the myo-inositol spin system when using a single spin-echo localization method we recently introduced. Glycine was detected at TE = 20 ms with an average Cramér-Rao lower bound (CRLB) of 8.6% +/- 1.5% in rat brain (N = 5), at 9.4 T. The concentration of glycine was determined using LCModel analysis at 1.1 +/- 0.1 mM, in good agreement with biochemical measurements previously reported. We conclude that at high magnetic fields, glycine can be measured at a relatively short echo time (TE) without additional editing efforts.

Non-invasive detection of cocaine dissolved in wine bottles by (1) H magnetic resonance spectroscopy.

Recently, a number of cases of smuggling dissolved cocaine in wine bottles have been reported. The aim of the present study was to determine whether cocaine dissolved in wine can be detected by proton magnetic resonance spectroscopy ((1) H MRS) on a standard clinical MR scanner, in intact (i.e. unopened) wine bottles. (1) H MRS experiments were performed with a 3 Tesla clinical scanner on wine phantoms with or without cocaine contamination. The aromatic protons of cocaine displayed resonance peaks in the 7-8 ppm region of the spectrum, where no overlapping resonances of wine were present. Additional cocaine resonances were detected in the 2-3 ppm region of the spectrum, between the resonances of ethanol and other wine constituents. Detection of cocaine in wine (at 5 mM, i.e. ∼1.5 g/L) was feasible in a scan time of 1 min. We conclude that dissolved cocaine can be detected in intact wine bottles, on a standard clinical MR scanner. Thus, (1) H MRS is the technique of choice to examine this type of suspicious cargo, since it allows for a non-destructive and rapid content characterization.

Occurrence and fate of micropollutants in the Vidy Bay of Lake Geneva, Switzerland. Part I: priority list for environmental risk assessment of pharmaceuticals.

Pharmaceuticals are substances designed to have a biological effect in humans. Their presence in the environment, especially in surface waters, is of increasing concern because of their potential risk to non-target species. A large number of pharmaceuticals are on the market; for example, approximately 2,000 active ingredients are approved in Europe, and many of them have already been detected in surface water. It is therefore crucial to select the substances that may do the most harm to the environment prior to performing measurements and extensive risk assessment. In the present study, a method to determine a list of pharmaceuticals to survey in surface water is proposed. Inclusion of substances on the list was based on a screening procedure, the analytical feasibility, and previous knowledge of pharmaceuticals detected in water. The screening procedure proposed here is an improvement on the standard procedure of the European Medicine Evaluation Agency (EMEA). It is designed to decrease the number of pharmaceuticals to be evaluated in a stepwise manner, thus decreasing the number of data necessary for the evaluation. We applied our approach to determine a list of 37 pharmaceuticals and four hormones to survey in a specific region of Switzerland, the Lake Geneva area, and discussed the advantages and weak points of the method.

Occurrence and fate of micropollutants in the Vidy Bay of Lake Geneva, Switzerland. Part II: micropollutant removal between wastewater and raw drinking water.

The occurrence and removal of 58 pharmaceuticals, endocrine disruptors, corrosion inhibitors, biocides, and pesticides, were assessed in the wastewater treatment plant (WWTP) of the city of Lausanne, Switzerland, as well as in the effluent-receiving water body, the Vidy Bay of Lake Geneva. An analytical screening method to simultaneously measure all of the 58 micropollutants was developed based on ultra performance liquid chromatography coupled to a tandem mass spectrometer (UPLC-MS/MS). The selection of pharmaceuticals was primarily based on a prioritization study, which designated them as environmentally relevant for the Lake Geneva region. Except for the endocrine disruptor 17alpha-ethinylestradiol, all substances were detected in 24-h composite samples of wastewater entering the WWTP or in the treated effluent. Of these compounds, 40% were also detected in raw drinking water, pumped from the lake 3 km downstream of the WWTP. The contributions of dilution and degradation to micropollutant elimination between the WWTP outlet and the raw drinking water intake were established in different model scenarios using hypothetical residence times of the wastewater in Vidy Bay of 1, 4, or 90 d. Concentration decrease due to processes other than dilution was observed for diclofenac, beta-blockers, several antibiotics, corrosion inhibitors, and pesticides. Measured environmental concentrations (MECs) of pharmaceuticals were compared to the predicted environmental concentrations (PECs) determined in the prioritization study and agreed within one order of magnitude, but MECs were typically greater than the corresponding PECs. Predicted no-effect concentrations of the analgesic paracetamol, and the two antibiotics ciprofloxacin and sulfamethoxazole, were exceeded in raw drinking water samples and therefore present a potential risk to the ecosystem.

Evidence of chemical exchange in recombinant Major Urinary Protein and quenching thereof upon pheromone binding.

The internal dynamics of recombinant Major Urinary Protein (rMUP) have been investigated by monitoring transverse nitrogen-15 relaxation using multiple-echo Carr-Purcell-Meiboom-Gill (CPMG) experiments. While the ligand-free protein (APO-rMUP) features extensive evidence of motions on the milliseconds time scale, the complex with 2-methoxy-3-isobutylpyrazine (HOLO-rMUP) appears to be much less mobile on this time scale. At 308 K, exchange rates k (ex) = 500-2000 s(-1) were typically observed in APO-rMUP for residues located adjacent to a beta-turn comprising residues 83-87. These residues occlude an entry to the binding pocket and have been proposed to be a portal for ligand entry in other members of the lipocalin family, such as the retinol binding protein and the human fatty-acid binding protein. Exchange rates and populations are largely uncorrelated, suggesting local 'breathing' motions rather than a concerted global conformational change.

Effects of protein-pheromone complexation on correlated chemical shift modulations.

Major urinary protein (MUP) is a pheromone-carrying protein of the lipocalin family. Previous studies by isothermal titration calorimetry (ITC) show that the affinity of MUP for the pheromone 2-methoxy-3-isobutylpyrazine (IBMP) is mainly driven by enthalpy, with a small unfavourable entropic contribution. Entropic terms can be attributed in part to changes in internal motions of the protein upon binding. Slow internal motions can lead to correlated or anti-correlated modulations of the isotropic chemical shifts of carbonyl C' and amide N nuclei. Correlated chemical shift modulations (CSM/CSM) in MUP have been determined by measuring differences of the transverse relaxation rates of zero- and double-quantum coherences ZQC{C'N} and DQC{C'N}, and by accounting for the effects of correlated fluctuations of dipole-dipole couplings (DD/DD) and chemical shift anisotropies (CSA/CSA). The latter can be predicted from tensor parameters of C' and N nuclei that have been determined in earlier work. The effects of complexation on slow time-scale protein dynamics can be determined by comparing the temperature dependence of the relaxation rates of APO-MUP (i.e., without ligand) and HOLO-MUP (i.e., with IBMP as a ligand).

Evidence of slow motions by cross-correlated chemical shift modulation in deuterated and protonated proteins.

Cross-correlated fluctuations of isotropic chemical shifts can provide evidence for slow motions in biomolecules. Slow side-chain dynamics have been investigated in (15)N and (13)C enriched ubiquitin by monitoring the relaxation of C(alpha)-C(beta) two-spin coherences (Frueh et al., 2001). This method, which had hitherto been demonstrated only for protonated ubiquitin, has now been applied to both protonated and deuterated proteins. Deuteration reduces the dipole-dipole contributions to the DD/DD cross-correlation, thus facilitating the observation of subtle effects due to cross-correlation of the fluctuations of the isotropic (13)C chemical shifts. The decays of double- and zero-quantum coherences are significantly slower in the deuterated protein than in the protonated sample. Slow motions are found both in loops and in secondary structure elements.

Thermodynamics of binding of 2-methoxy-3-isopropylpyrazine and 2-methoxy-3-isobutylpyrazine to the major urinary protein.

In the present study we examine the thermodynamics of binding of two related pyrazine-derived ligands to the major urinary protein, MUP-I, using a combination of isothermal titration calorimetry (ITC), X-ray crystallography, and NMR backbone (15)N and methyl side-chain (2)H relaxation measurements. Global thermodynamics data derived from ITC indicate that binding is driven by favorable enthalpic contributions, rather than the classical entropy-driven hydrophobic effect. Unfavorable entropic contributions from the protein backbone and side-chain residues in the vicinity of the binding pocket are partially offset by favorable entropic contributions at adjacent positions, suggesting a "conformational relay" mechanism whereby increased rigidity of residues on ligand binding are accompanied by increased conformational freedom of side chains in adjacent positions. The principal driving force governing ligand affinity and specificity can be attributed to solvent-driven enthalpic effects from desolvation of the protein binding pocket.