Proton Relaxometry Research Articles

Proton relaxometry of tree leaves at hypogeomagnetic fields.

We report on a cross-species proton-relaxometry study in ex vivo tree leaves using nuclear magnetic resonance (NMR) at 7µT. Apart from the intrinsic interest of probing nuclear-spin relaxation in biological tissues at magnetic fields below Earth field, our setup enables comparative analysis of plant water dynamics without the use of expensive commercial spectrometers. In this work, we focus on leaves from common Eurasian evergreen and deciduous tree families: Pinaceae (pine, spruce), Taxaceae (yew), Betulaceae (hazel), Prunus (cherry), and Fagaceae (beech, oak). Using a nondestructive protocol, we measure their effective proton T 2 relaxation times as well as track the evolution of water content associated with leaf dehydration. Newly developed "gradiometric quadrature" detection and data-processing techniques are applied in order to increase the signal-to-noise ratio (SNR) of the relatively weak measured signals. We find that while measured relaxation times do not vary significantly among tree genera, they tend to increase as leaves dehydrate. Such experimental modalities may have particular relevance for future drought-stress research in ecology, agriculture, and space exploration.

Inorganic Phosphate-Induced Extracellular Vesicles from Vascular Smooth Muscle Cells Contain Elevated Levels of Hyaluronic Acid, Which Enhance Their Interaction with Very Small Superparamagnetic Iron Oxide Particles.

Patients with chronic kidney disease (CKD) have a high prevalence of hyperphosphatemia, where uremic toxins like inorganic phosphate (Pi) induce a cardiovascular remodeling. Related disorders like atherosclerosis bear the risk of increased morbidity and mortality. We previously found that Pi stimulates the synthesis and sulfation of the negatively charged glycosaminoglycans (GAGs) heparan sulfate and chondroitin sulfate in vascular smooth muscle cells (VSMC). Similar GAG alterations were detected in VSMC-derived exosome-like extracellular vesicles (EV). These EV showed a strong interaction with very small superparamagnetic iron oxide particles (VSOP), which are used as imaging probes for experimental magnetic resonance imaging (MRI). Hyaluronic acid (HA) represents another negatively charged GAG which is supposed to function as binding motif for VSOP as well. We investigated the effects of Pi on the amounts of HA in cells and EV and studied the HA-dependent interaction between VSOP with cells and EV. Rat VSMC were treated with elevated concentrations of Pi. CKD in rats was induced by adenine feeding. EV were isolated from culture supernatants and rat plasma. We investigated the role of HA in binding VSOP to cells and EV via cell-binding studies, proton relaxometry, and analysis of cellular signaling, genes, proteins, and HA contents. Due to elevated HA contents, VSMC and EV showed an increased interaction with VSOP after Pi stimulation. Amongst others, Pi induced hyaluronan synthase (HAS)2 expression and activation of the Wnt pathway in VSMC. An alternative upregulation of HA by iloprost and an siRNA-mediated knockdown of HAS2 confirmed the importance of HA in cells and EV for VSOP binding. The in vitro-derived data were validated by analyses of plasma-derived EV from uremic rats. In conclusion, the inorganic uremic toxin Pi induces HA synthesis in cells and EV, which leads to an increased interaction with VSOP. HA might therefore be a potential molecular target structure for improved detection of pathologic tissue changes secondary to CKD like atherosclerosis or cardiomyopathy using EV, VSOP and MRI.

New milk protein anisotropic structures formed by high moisture extrusion

In this work, the formation of anisotropic, layered structures from milk protein concentrates was investigated using high moisture extrusion. These structures were obtained in combination with three concentrations of starch (0 %, 14 %, and 33 %, d.b), and at three different moisture levels: 58 %, 63 %, and 68 %. Two different temperature profiles were investigated, reaching a maximum barrel temperature of 80 and 115 °C (and maximum melt temperature of 56 and 64 °C, respectively), with a constant screw speed and feed rate. In situ monitoring of torque, as well as the evaluation of water partitioning via proton relaxometry highlighted the interplay between moisture content and barrel temperature in starch transitions and water intake. For example, the presence of starch displayed water populations of distinct mobility, featured by higher relaxation times, and the apparition (or intensification) of new peaks representing free water in the system. The extruded samples showed a multi-phase system with swollen starch granules embedded in a protein network when 14 % starch was used, at moisture <63 %, as depicted by confocal microscopy. Tensile strength and cutting force tests showed a broad range of textures, from more brittle at high starch concentration, to softer and mushier when extruded at high moisture. Increasing starch and moisture content disrupted the protein network structure. This work demonstrated that milk protein concentrates show great potential to create novel anisotropic structures by extrusion, with tunable texture properties depending on starch, moisture and temperature conditions.

Gadolinium-Doped Carbon Nanoparticles with Red Fluorescence and Enhanced Proton Relaxivity as Bimodal Nanoprobes for Bioimaging Applications

Carbon-based nanoparticles (CNPs) have demonstrated great potential in biomedical applications because of their unique physical and chemical properties, and excellent biocompatibility. Herein, we have studied two types of CNPs with gadolinium (Gd) impurities (Gd-CNPs), which were prepared by microwave synthesis (MWS) and hydrothermal synthesis (HTS), for potential applications as photoluminescent (PL) labels and contrast agents in magnetic resonance imaging (MRI). The prepared Gd-CNPs were investigated by means of transmission electron microscopy (TEM), Fourier-transform infrared spectroscopy, UV–visible absorption spectroscopy, and magnetic-resonance relaxometry, which allowed us to reveal specific features and functional properties of the prepared samples. While the TEM data showed similar size distributions of both types of Gd-CNPs with mean sizes of 4–5 nm, the optical absorption spectroscopy showed higher absorption in the visible spectral region and stronger PL in the red and near-infrared (NIR) spectral regions for the MWS samples in comparison with those prepared by HTS. Under green light excitation the former samples exhibited the bright red-NIR PL with quantum efficiency of the order of 10%. The proton relaxometry measurements demonstrated that the HTS samples possessed longitudinal and transverse relaxivities of about 42 and 70 mM−1s−1, whereas the corresponding values for the MWS samples were about 8 and 13 mM−1s−1, respectively. The obtained results can be useful for the selection of appropriate synthesis conditions for carbon-based nanoparticles for bimodal bioimaging applications.

Incorporation of PEG Diacrylates (PEGDA) Generates Hybrid Fmoc-FF Hydrogel Matrices.

Generated by a hierarchical and multiscale self-assembling phenomenon, peptide-based hydrogels (HGs) are soft materials useful for a variety of applications. Short and ultra-short peptides are intriguing building blocks for hydrogel fabrication. These matrices can also be obtained by mixing low-molecular-weight peptides with other chemical entities (e.g., polymers, other peptides). The combination of two or more constituents opens the door to the development of hybrid systems with tunable mechanical properties and unexpected biofunctionalities or morphologies. For this scope, the formulation, the multiscale analysis, and the supramolecular characterization of novel hybrid peptide-polymer hydrogels are herein described. The proposed matrices contain the Fmoc-FF (Nα-fluorenylmethyloxycarbonyl diphenylalanine) hydrogelator at a concentration of 0.5 wt% (5.0 mg/mL) and a diacrylate α-/ω-substituted polyethylene-glycol derivative (PEGDA). Two PEGDA derivatives, PEGDA 1 and PEGDA2 (mean molecular weights of 575 and 250 Da, respectively), are mixed with Fmoc-FF at different ratios (Fmoc-FF/PEGDA at 1/1, 1/2, 1/5, 1/10 mol/mol). All the multicomponent hybrid peptide-polymer hydrogels are scrutinized with a large panel of analytical techniques (including proton relaxometry, FTIR, WAXS, rheometry, and scanning electronic microscopy). The matrices were found to be able to generate mechanical responses in the 2-8 kPa range, producing a panel of tunable materials with the same chemical composition. The release of a model drug (Naphthol Yellow S) is reported too. The tunable features, the different topologies, and the versatility of the proposed materials open the door to the development of tools for different applicative areas, including diagnostics, liquid biopsies and responsive materials. The incorporation of a diacrylate function also suggests the possible development of interpenetrating networks upon cross-linking reactions. All the collected data allow a mutual comparison between the different matrices, thus confirming the significance of the hybrid peptide/polymer-based methodology as a strategy for the design of innovative materials.

How the cation size impacts on the relaxational and diffusional dynamics of supercooled butylammonium-based ionic liquids: DPEBA–TFSI versus BTMA–TFSI

Abstract Li-bis(trifluoromethylsulfonyl)imide based ionic liquids with either butyl-trimethylammonium or N,N-dimethyl-N-(2-(propionyloxy)-ethyl)butan-1-ammonium as the anion were studied using proton and fluorine relaxometry as well as using field-gradient diffusometry to gain separate access to cation and anion dynamics in these compounds. The transport parameters obtained for these ionic liquids are compared with the estimates based on the conductivity data from literature and from the present work. The impact of cation size on correlation effects, the latter parameterized in terms of various Haven ratios, is mapped out.

Mn2+ Complexes with Pyclen-Based Derivatives as Contrast Agents for Magnetic Resonance Imaging: Synthesis and Relaxometry Characterization.

Magnetic resonance imaging (MRI) has a leading place in medicine as an imaging tool of high resolution for anatomical studies and diagnosis of diseases, in particular for soft tissues that cannot be accessible by other modalities. Many research works are thus focused on improving the images obtained with MRI. This technique has indeed poor sensitivity, which can be compensated by using a contrast agent (CA). Today, the clinically approved CAs on market are solely based on gadolinium complexes that may induce nephrogenic systemic fibrosis for patients with kidney failure, whereas more recent studies on healthy rats also showed Gd retention in the brain. Consequently, researchers try to elaborate other types of safer MRI CAs like manganese-based complexes. In this context, the synthesis of Mn2+ complexes of four 12-membered pyridine-containing macrocyclic ligands based on the pyclen core was accomplished and described herein. Then, the properties of these Mn(II) complexes were studied by two relaxometric methods, 17O NMR spectroscopy and 1H NMR dispersion profiles. The time of residence (τM) and the number of water molecules (q) present in the inner sphere of coordination were determined by these two experiments. The efficacy of the pyclen-based Mn(II) complexes as MRI CAs was evaluated by proton relaxometry at a magnetic field intensity of 1.41 T near those of most medical MRI scanners (1.5 T). Both the 17O NMR and the nuclear magnetic relaxation dispersion profiles indicated that the four hexadentate ligands prepared herein left one vacant coordination site to accommodate one water molecule, rapidly exchanging, in around 6 ns. Furthermore, it has been shown that the presence of an additional amide bond formed when the paramagnetic complex is conjugated to a molecule of interest does not alter the inner sphere of coordination of Mn, which remains monohydrated. These complexes exhibit r1 relaxivities, large enough to be used as clinical MRI CAs (1.7-3.4 mM-1·s-1, at 1.41 T and 37 °C).

Nondestructive measurement of intramuscular fat content of fresh beef meat by a hand-held magnetic resonance sensor

ABSTRACT To realize nondestructive in-situ quantification of intramuscular fat in fresh beef meat, a lightweight (5.0-kg) hand-held sensor that consists of a planar radio-frequency coil and a unilateral (i.e., single-sided) magnetic circuit was constructed as a subunit of a time-domain proton magnetic resonance (MR) scanner system. The MR scanner based on proton relaxometry enables the nondestructive quantification of the fat content of meat by using the difference in the spin-spin relaxation times (T2) of water molecules in muscle and of fat molecules. The investigation depth of the sensor unit was 7 mm, which is sufficient to probe the meat beneath a subcutaneous fat layer less than a few millimeters thick. The scanner was successfully applied in a laboratory at a meat temperature of 30.5°C to quantify the fat content at 30 locations in fresh beef samples. The required measurement time was 22 s for each location. Reasonable agreement with a root-mean-square error as small as 5.4 wt% was obtained for fat quantification compared with the conventional destructive food analysis (Soxhlet extraction method). Thus, the portable MR scanner with a hand-held sensor unit is a promising nondestructive and noninvasive tool for the in-situ fat quantification of fresh beef (e.g., carcasses) with thin subcutaneous fat layers at meat processing factories.

Effect of modified microcrystalline cellulose on poly(3-hydroxybutyrate) molecular dynamics by proton relaxometry

Hybrid films of poly(3-hydroxybutyrate) (PHB) and modified cellulose by solution casting method were prepared, aiming to study the influence of modified cellulose (0.25%, 0.5% and 0.75% w/w) on the structural organization and thermal properties of PHB matrix. The modified cellulose showed good dispersion in polymer matrix, due to the high compatibility between phases explained by similarity of polymer and filler structures. The composites were studied by wide-angle X-ray diffractometry, differential scanning calorimetry, thermogravimetric analysis, and time-domain nuclear magnetic resonance (NMR) spectroscopy. The conjugation of results coming from those techniques allowed to determine the cellulose concentration that had the highest influence on crystalline planes and degree of crystallinity of PHB, that is, the influence in the composite structure. The molecular dynamics obtained by NMR showed a reciprocal influence between polymer matrix and cellulose particles, enhancing the interactions present in the agglomerated particles.

Proton Relaxometry of Long-Lived Spin Order.

A study of long-lived spin order in chlorothiophene carboxylates at both high and low magnetic fields is presented. Careful sample preparation (removal of dissolved oxygen in solution, chelating of paramagnetic impurities, reduction of convection) allows one to obtain very long-lived singlet order of the two coupled protons in chlorothiophene derivatives, having lifetimes of about 130 s in D2 O and 240 s in deuterated methanol, which are much longer than the T1 -relaxation times (18 and 30 s, respectively, at a field =9.4 T). In protonated solvents the relaxation times become shorter, but the lifetime is still substantially longer than . In addition, long-lived coherences are shown to have lifetimes as long as 30 s. Thiophene derivatives can be used as molecular tags to study slow transport, slow dynamics and slow chemical processes, as has been shown in recent years.

Real‐time monitoring by proton relaxometry of radical polymerization reactions of acrylamide in aqueous solution

AbstractThe potential of time‐domain nuclear magnetic resonance (TD‐NMR) for the real‐time monitoring of solution radical polymerizations is demonstrated. A model system composed of a redox‐pair initiator system, acrylamide as monomer and water as solvent was investigated. A second‐generation continuous wave free precession technique was employed to measure the longitudinal relaxation time constant (T1) of the samples throughout the polymerization reactions. This parameter was shown to be sensitive to the reactant feed free‐radical enhancement of the water molecule relaxation time, making it a good probe to monitor monomer conversion in real time in an automated, non‐destructive fashion. It was found that the T1 value was better than the transverse relaxation time constant (T2) for describing the evolution of the polymerization reactions, due to its greater sensitivity to paramagnetic effects. The TD‐NMR signal variation observed was linked to the formation, propagation and termination steps of the radical polymerization kinetics scheme. These first results may contribute to the application of real‐time monitoring of radical polymerization reactions employing low‐cost and robust TD‐NMR spectrometers. © 2018 Society of Chemical Industry

Hybrid membranes of sulfonated poly ether ether ketone, ionic liquid and organically modified montmorillonite for proton exchange membranes with enhanced ionic conductivity and ionic liquid lixiviation protection

Through the fabrication of hybrid membranes, composed by the protic ionic liquid diethylmethylammonium trifluomethane sulfonate (demaTfO), sulfonated poly (ether ether kethone) with degree of sulfonation 73% (sPEEK73) and montmorillonite modified with dema+ cation (Mmtdema), this work evaluates the effect exerted by the compatibilization between clay and ionic liquid on the morphology and performance of membranes, as a function of Mmtdema proportion. This system of membranes showed high conductivity (78 mS/cm at 70°C) with some issues, due to the plasticization effect caused by montmorillonite and ionic liquid; these effects were confirmed through electrochemical impedance spectrometry (EIS), dynamic mechanical analysis (DMA) and proton relaxometry. The membranes showed resistance to ionic liquid lixiviation, which was confirmed through static water immersion and this effect was attributed to the protection effect that interlamellar spaces can provide to the ionic liquid, as it was visualized through TEM and XRD. The conjugation of results from several different techniques as SAXS, EIS, NMR, TEM and XRD permitted to obtain evidences for a proposed descriptive model of proton transporting process, which turns out to be a synergy between Grotthuss mechanism and vehicular mechanism, favored by the structure of Mmtdema and the preparation method, leading thus to high conductivity membranes with lixiviation protection for application as proton exchange membranes.

The synthesis of doxorubicin-conjugated magnetite nanoparticles and their magnetic resonance and cytotoxic properties

The possibility of increasing the effectiveness of antitumor drugs such as doxorubicin by preparing its complex with ultrafine magnetic iron oxide nanoparticles is considered. A method for binding doxorubicin molecules to magnetic nanoparticles via citric acid is proposed. The main magnetic properties of the obtained conjugates were studied by proton relaxometry and Mossbauer spectroscopy, while their cytotoxic activity was evaluated via spectrophotometric MTT assay in HeLa cells. It was shown that the conjugates of magnetite nanoparticles with doxorubicin are characterized by a high level of contrast in magnetic resonance imaging. The magnetic properties of doxorubicin-free and bound magnetite nanoparticles are mainly determined by the average size of nanoobjects and the phase composition and slightly depend on the composition of the stabilizing shell. The cytotoxic effect of the synthesized conjugates of magnetite nanoparticles with doxorubicin is higher than that of unbound doxorubicin. This makes it possible to increase the antitumor effect of doxorubicin and control the dynamics of its delivery in the form of a conjugate into the disease focus due to the magnetic contrast properties of nanoparticles.

The Effect of Montmorillonite Clay on the Crystallinity of Poly(vinyl alcohol) Nanocomposites Obtained by Solution Intercalation and In Situ Polymerization.

In this study, PVAL/sodic clay nanocomposite materials were obtained using different clay ratios. The nanocomposites were prepared by solution and in situ polymerization. They were characterized through proton relaxometry and X-ray diffraction, the latter technique used to determine the crystallinity of the PVAL matrix in the nanocomposites through the addition of unmodified montmorillonite clay. The relaxation data for the nanocomposite films obtained by solution intercalation showed a decrease in the proton spin-lattice relaxation values with increased clay proportion, forming a nanostructured material with high exfoliation degree, as a result of two phenomena: the higher molecular mobility of the polymer chains around the clay layers and the presence of paramagnetic metals in the clay layers, which accelerates the hydrogen relaxation process, causing a decrease in the relaxation value. The samples obtained through in situ polymerization were mixed nanocomposites (partly exfoliated and partly intercalated). The NMR relaxation data for the systems analyzed showed the solution intercalation process was more efficient than in situ polymerization in relation to the generation of nanostructured polymers with higher degree of exfoliated clay.

Solvent Effect on the Morphology of Lamellar Nanocomposites Based on HIPS

In this study, hybrid nanocomposites based on HIPS, high-impact polystyrene, were prepared in chloroform and in a 1:1 mixture of toluene and chloroform varying the quantity of organophilic montmorillonite. The morphology of the hybrid materials was investigated by X-ray diffraction (XRD) and proton relaxometry, determined in a low-field 1H nuclear magnetic resonance (NMR). The thermal characteristics were also obtained through thermogravimetric analysis. The emphasis of this study was to compare the effect of a pair of solvents against a single solvent in the formation/organization of the nanostructured materials when the solution method is applied. It was taken into consideration Interactions involving polymer-solvent, nanoparticle-solvent, and, principally, polymer-nanoparticle-solvent. The results show that the choice of solvent affects greatly the characteristic of the nanocomposites. Nanocomposites prepared with chloroform/toluene were more homogenous, more prone to exfoliate (for all organoclay concentrations), and had higher degradation temperature compared to the materials obtained with chloroform. It is clear from the data that the use of a solvent pair is important to produce good nanocomposites when using the solution method and that the clay proportion plays a role in the dispersion of organoclay in the polymer matrix.

Non-cooperative immobilization of residual water bound in lyophilized photosynthetic lamellae.

This study applied 1H-NMR in time and in frequency domain measurements to monitor the changes that occur in bound water dynamics at decreased temperature and with increased hydration level in lyophilizates of native wheat photosynthetic lamellae and in photosynthetic lamellae reconstituted from lyophilizate. Proton relaxometry (measured as free induction decay = FID) distinguishes a Gaussian component S within the NMR signal (o). This comes from protons of the solid matrix of the lamellae and consists of (i) an exponentially decaying contribution L1 from mobile membrane protons, presumably from lipids, and from water that is tightly bound to the membrane surface and thus restricted in mobility; and (ii) an exponentially decaying component L2 from more mobile, loosely bound water pool. Both proton relaxometry data and proton spectroscopy show that dry lyophilizate incubated in dry air, i.e., at a relative humidity (p/p0) of 0% reveals a relatively high hydration level. The observed liquid signal most likely originates from mobile membrane protons and a tightly bound water fraction that is sealed in pores of dry lyophilizate and thus restricted in mobility. The estimations suggest that the amount of sealed water does not exceed the value characteristic for the main hydration shell of a phospholipid. Proton spectra collected for dry lyophilizate of photosynthetic lamellae show a continuous decrease in the liquid signal component without a distinct freezing transition when it is cooled down to -60ºC, which is significantly lower than the homogeneous ice nucleation temperature [Bronshteyn, V.L. et al. Biophys. J. 65 (1993) 1853].

Mössbauer spectroscopy and proton relaxometry study of magnetite nanoparticles designed for preparing diagnostic contrast media

Mossbauer spectroscopy, proton relaxometry, and transmission electron microscopy are used to study magnetite nanoparticles designed for creating diagnostic contrast media. Superparamagnetic magnetite nanoparticles with a size of 5–7 nm and blocking temperature of T b = 50 K are examined as a component of diagnostic contrast media with relaxation times T 1 and T 2 capable of circulating in the bloodstream for a long time. Larger ferrimagnetic nanoparticles (30–40 nm) can be concentrated in pathological tissues by applying an external magnetic field, thereby providing a means for hyperthermia.

The influence of NaYF₄:Yb,Er size/phase on the multimodality of co-encapsulated magnetic photon-upconverting polymeric nanoparticles.

We report the synthesis, characterisation and evaluation of the in vitro biocompatibility of polymeric nanoparticles with both magnetic and upconverting fluorescent properties. The particles consist of superparamagnetic iron oxide nanoparticles and upconverting NaYF4:Yb,Er nanoparticles co-encapsulated within a poly(glycidyl methacrylate) sphere. Two different upconverting nanoparticles (10 nm α-NaYF4:Yb,Er and 50 nm β-NaYF4:Yb,Er) were synthesised and the optical and magnetic properties of the composite polymeric nanoparticle systems assessed by near infra-red laser spectroscopy, SQUID magnetometry and proton relaxometry. A live-dead assay was used to assess the viability of PC-12 neural cells incubated with varying concentrations of the nanoparticles. The composite nanoparticles produced no observed impact on cellular viability even at concentrations as high as 1000 μg mL(-1). Confocal microscopy revealed uptake of nanoparticles by PC-12 cells and peri-nuclear cytoplasmic localisation. Both particle systems show favourable magnetic properties. However, only the nanospheres containing 50 nm β-NaYF4:Yb,Er were suitable for optical tracking because the presence of iron oxide within the composites imparts a significant quenching of the upconversion emission. This study demonstrates the size and phase of the upconverting nanoparticles are important parameters that have to be taken into account in the design of multimodal nanoparticles using co-encapsulation strategies.

1H NMR relaxometry and X-ray study of PCL/nevirapine hybrids

Polycaprolactone (PCL) hybrids containing layered silicates, silica and dioxide titanium were developed and used as matrixes for the incorporation of nevirapine. These systems were characterized by X-ray diffraction (XRD) and proton nuclear magnetic resonance (NMR) relaxometry. The XRD showed that the PCL crystallization is hindered with higher amounts of nevirapine and that the drug has an amorphous distribution in the PCL matrix. The NMR showed that the hybrids present three distinct environments: crystallites, rigid-amorphous and flexible-amorphous regions, with three distinct spin-lattice relaxation times, T11, T12, T13. The proton relaxometry showed that the PCL chains have more restricted motions in the presence of Viscogel organoclay S7 comparing to TiO2 and SiO2. The presence of nevirapine and nanoparticles together decreased the polymer chain mobility, especially in the amorphous regions. Both T12 and T13 dispersions could be interpreted in terms of power laws T12, T13 ∼ νp. In the case of T12, the exponent obtained was around 0.73, which in the frame of the renormalized Rouse model could be associated with a type I mesophase region, indicating some constraint in the amorphous part.

Initial Phases of Antarctic Ramalina terebrata Hook f. &amp; Taylor Thalli Rehydration Observed by Proton Relaxometry

Hydration kinetics, sorption isotherm, and proton free induction decays are measured for Ramalina terebrata thalli rehydrated from gaseous phase. Very tightly, tightly, and loosely bound water fractions are distinguished. Sorption isotherm is sigmoidal in form with the mass of water saturating primary water binding sites equal to ∆m/m0 = 0.046. Proton free induction decays show the presence of immobilized water fraction (T ∗ 2L1 ≈ 100 μs) and mobile water pool (T ∗ 2L2 ≈ 330 μs). Sorption isotherm fitted to the NMR data shows the absence of water fraction “sealed” in pores of dry thallus.