39 research outputs found
Investigation of quorum sensing in Candida albicans
The dimorphic fungus Candida albicans produces extracellular farnesol which acts as a quorum-sensing molecule (QSM) to suppress filamentation. Of four possible geometric isomers of farnesol, only the E,E isomer possesses QSM activity. We tested number of natural and synthetic analogs of farnesol for their activity in an N-acetylglucosamine-induced differentiation assay for germ tube formation (GTF). Modified structural features include the nature of the head group, chain length, presence or absence of the three double bonds, substitution of a backbone carbon by S, O, N, and Se heteroatoms, presence or absence of a 3-methyl branch, as well as the bulkiness and the hydrophobic tail. In order to elucidate localization a putative receptor we prepared ten polyenes typified by 3,7,11-trimethyl-2,4,6,8,10-dodecapentenaldehyde oxime. Four of the ten analogs display strong quorum-sensing activity in the human pathogen Candida albicans, also they are fluorescent. Therefore oxime anti-4 is demonstrated to be useful for confocal fluorescence microscopic imaging of fungal cells. Also, we have investigated of a series of farnesol analogs replacing the primary alcohol head group with several classes of heterocycles. High levels of quorum-sensing activity were associated with several types of heterocycles. Two non toxic analogs were tested in C. albicans inoculated immune-challenged mice. One of the two analogs has been shown to perfectly emulate farnesol in vivo behavior, which unfortunately includes an increase of C. Albicans virulence in the mice. In an attempt to isolate putative farnesol binding receptor we synthesized of affinity media designed to isolate a putative farnesol receptor or binding protein in C. albicans. The approach rests on a new class of farnesol analogs, which maintain the acidic head group and farnesyl-like backbone known from our earlier studies to be necessary for quorum sensing, but which also allow for covalent chemical linkage of the head group to an affinity support. In preliminary results, affinity chromatography of the total protein extract of C. albicans resulted in selective retention of a handful of proteins. Mass spectrometry of some of eluted proteins reveals two matches against known fungal proteins
Spin–Lattice Relaxation of Hyperpolarized Metronidazole in Signal Amplification by Reversible Exchange in Micro-Tesla Fields
Simultaneous
reversible chemical exchange of <i>para</i>-hydrogen and
to-be-hyperpolarized substrate on metal centers enables
spontaneous transfer of spin order from <i>para</i>-hydrogen
singlet to nuclear spins of the substrate. When performed at a sub-micro-tesla
magnetic field, this technique of NMR signal amplification by reversible
exchange in shield enables alignment transfer to heteronuclei (SABRE-SHEATH).
SABRE-SHEATH has been shown to hyperpolarize nitrogen-15 sites of
a wide range of biologically interesting molecules to a high polarization
level (<i>P</i> > 20%) in 1 min. Here, we report on a
systematic
study of <sup>1</sup>H, <sup>13</sup>C, and <sup>15</sup>N spin–lattice
relaxation (<i>T</i><sub>1</sub>) of metronidazole-<sup>13</sup>C<sub>2</sub>-<sup>15</sup>N<sub>2</sub> in the SABRE-SHEATH
hyperpolarization process. In the micro-tesla range, we find that
all <sup>1</sup>H, <sup>13</sup>C, and <sup>15</sup>N spins studied
share approximately the same <i>T</i><sub>1</sub> values
(ca. 4 s under the conditions studied) because of mixing of their
Zeeman levels, which is consistent with the model of relayed SABRE-SHEATH
effect. These <i>T</i><sub>1</sub> values are significantly
lower than those at a higher magnetic field (i.e. the Earth’s
magnetic field and above), which exceed 3 min in some cases. Moreover,
these relatively short <i>T</i><sub>1</sub> values observed
below 1 μT limit the polarization build-up process of SABRE-SHEATH,
thereby limiting the maximum attainable <sup>15</sup>N polarization.
The relatively short <i>T</i><sub>1</sub> values observed
below 1 μT are primarily caused by intermolecular interactions
with quadrupolar iridium centers or dihydride protons of the employed
polarization transfer catalyst, whereas intramolecular spin–spin
interactions with <sup>14</sup>N quadrupolar centers have a significantly
smaller contribution. The presented experimental results and their
analysis will be beneficial for more rational design of SABRE-SHEATH
(i) polarization transfer catalysts and (ii) hyperpolarized molecular
probes in the context of biomedical imaging and other applications
Parahydrogen-Induced Polarization with a Rh-Based Monodentate Ligand in Water
Reported here is a water-soluble RhÂ(I)-based catalyst
for performing
parahydrogen-induced polarization (PHIP). The [RhÂ(I)Â(norbornadiene)Â(THP)<sub>2</sub>]<sup>+</sup>[BF<sub>4</sub>]<sup>−</sup> catalyst
utilizes the monodentate phosphine ligand trisÂ(hydroxymethyl)Âphosphine
(THP). The monodentate PHIP catalyst is less susceptible to oxygenation
by air and the THP ligand is significantly less expensive than bidentate
water-soluble PHIP ligands. In situ PHIP detection with this monodentate
RhÂ(I)-based catalyst in water yielded 12% <sup>13</sup>C polarization
for the parahydrogen addition product, 2-hydroxyethyl 1-<sup>13</sup>C-propionate-d<sub>2,3,3</sub> (HEP), with a <sup>13</sup>C <i>T</i><sub>1</sub> relaxation of 108 s at 0.0475 T. PHIP polarization
yields were high, reflecting efficient hydrogenation even under conditions
of high content of the oxidized phosphine form of the THP ligand
Efficient Synthesis of Nicotinamide-1-<sup>15</sup>N for Ultrafast NMR Hyperpolarization Using Parahydrogen
Nicotinamide (a vitamin B<sub>3</sub> amide) is one of the key
vitamins as well as a drug for treatment of M. tuberculosis, HIV, cancer, and other diseases. Here, an improved Zincke reaction
methodology is presented allowing for straightforward and scalable
synthesis of nicotinamide-1-<sup>15</sup>N with an excellent isotopic
purity (98%) and good yield (55%). <sup>15</sup>N nuclear spin label
in nicotinamide-1-<sup>15</sup>N can be NMR hyperpolarized in seconds
using parahydrogen gas. NMR hyperpolarization using the process of
temporary conjugation between parahydrogen and to-be-hyperpolarized
biomolecule on hexacoordinate iridium complex via the Signal Amplification
By Reversible Exchange (SABRE) method significantly increases detection
sensitivity (e.g., >20 000-fold for nicotinamide-1-<sup>15</sup>N at 9.4 T) as has been shown by Theis T. et al. (<i>J. Am.
Chem. Soc.</i> <b>2015</b>, <i>137</i>, 1404),
and hyperpolarized in this fashion, nicotinamide-1-<sup>15</sup>N
can be potentially used to probe metabolic processes in vivo in future
studies. Moreover, the presented synthetic methodology utilizes mild
reaction conditions, and therefore can also be potentially applied
to synthesis of a wide range of <sup>15</sup>N-enriched N-heterocycles
that can be used as hyperpolarized contrast agents for future in vivo
molecular imaging studies
Parahydrogen Induced Polarization of 1-<sup>13</sup>C‑Phospholactate‑<i>d</i><sub>2</sub> for Biomedical Imaging with >30,000,000-fold NMR Signal Enhancement in Water
The synthetic protocol for preparation
of 1-<sup>13</sup>C-phosphoenolpyruvate-<i>d</i><sub>2</sub>, precursor for parahydrogen-induced polarization
(PHIP) of 1-<sup>13</sup>C-phospholactate-<i>d</i><sub>2</sub>, is reported. <sup>13</sup>C nuclear spin polarization of 1-<sup>13</sup>C-phospholactate-<i>d</i><sub>2</sub> was increased
by >30,000,000-fold (5.75 mT) in water. The reported <sup>13</sup>C polarization level approaching unity (>15.6%), long lifetime
of <sup>13</sup>C hyperpolarized 1-<sup>13</sup>C-phospholactate-<i>d</i><sub>2</sub> (58 ± 4 s versus 36 ± 2 s for nondeuterated
form at 47.5 mT), and large production quantities (52 μmoles
in 3 mL) in aqueous medium make this compound useful as a potential
contrast agent for the molecular imaging of metabolism and other applications
Synthesis of Unsaturated Precursors for Parahydrogen-Induced Polarization and Molecular Imaging of 1-<sup>13</sup>C‑Acetates and 1-<sup>13</sup>C‑Pyruvates via Side Arm Hydrogenation
Hyperpolarized forms of 1-<sup>13</sup>C-acetates and 1-<sup>13</sup>C-pyruvates are used as diagnostic
contrast agents for molecular
imaging of many diseases and disorders. Here, we report the synthetic
preparation of 1-<sup>13</sup>C isotopically enriched and pure from
solvent acetates and pyruvates derivatized with unsaturated ester
moiety. The reported unsaturated precursors can be employed for NMR
hyperpolarization of 1-<sup>13</sup>C-acetates and 1-<sup>13</sup>C-pyruvates via parahydrogen-induced polarization (PHIP). In this
PHIP variant, Side arm hydrogenation (SAH) of unsaturated ester moiety
is followed by the polarization transfer from nascent parahydrogen
protons to <sup>13</sup>C nucleus via magnetic field cycling procedure
to achieve hyperpolarization of <sup>13</sup>C nuclear spins. This
work reports the synthesis of PHIP-SAH precursors: vinyl 1-<sup>13</sup>C-acetate (55% yield), allyl 1-<sup>13</sup>C-acetate (70% yield),
propargyl 1-<sup>13</sup>C-acetate (45% yield), allyl 1-<sup>13</sup>C-pyruvate (60% yield), and propargyl 1-<sup>13</sup>C-pyruvate (35%
yield). Feasibility of PHIP-SAH <sup>13</sup>C hyperpolarization was
verified by <sup>13</sup>C NMR spectroscopy: hyperpolarized allyl
1-<sup>13</sup>C-pyruvate was produced from propargyl 1-<sup>13</sup>C-pyruvate with <sup>13</sup>C polarization of ∼3.2% in CD<sub>3</sub>OD and ∼0.7% in D<sub>2</sub>O. <sup>13</sup>C magnetic
resonance imaging is demonstrated with hyperpolarized 1-<sup>13</sup>C-pyruvate in aqueous medium
Parahydrogen-induced polarization of 14N nuclei
Hyperpolarization techniques provide a dramatic increase in sensitivity of nuclear magnetic resonance spectroscopy and imaging. In spite of the outstanding progress in solution-state hyperpolarization of spin-1/2 nuclei, hyperpolarization of quadrupolar nuclei remains challenging. Here, hyperpolarization of quadrupolar 14N nuclei with natural isotopic abundance of >99% is demonstrated. This is achieved via pairwise addition of parahydrogen to tetraalkylammonium salts with vinyl or allyl unsaturated moieties followed by a subsequent polarization transfer from 1H to 14N nuclei at high magnetic field using PH-INEPT or PH-INEPT+ radiofrequency pulse sequence. Catalyst screening identified water-soluble rhodium complex [Rh(P(m-C6H4SO3Na)3)3Cl] as the most efficient catalyst for hyperpolarization of the substrates under study, providing up to 1.3% and up to 6.6% 1H polarization in the cases of vinyl and allyl precursors, respectively. The performance of PH-INEPT and PH-INEPT+ pulse sequences was optimized with respect to interpulse delays, and the resultant experimental dependences were in good agreement with simulations. As a result, 14N NMR signal enhancement of up to 760-fold at 7.05 T (corresponding to 0.15% 14N polarization) was obtained