C. Andrea Boicelli

Phospholipid-Based Reverse Micelles

Physicochemical investigations on the aggregation of phospholipids (mainly phosphatidylcholines) in organic solvents are reviewed and compared with the aggregation behaviour of phospholipids in aqueous medium. In particular we review the data showing that phosphatidylcholines (lecithins) form reverse micellar structures in certain apolar solvents. In these systems not only low molecular weight compounds but also catalytically active enzymes and entire cells can be solubilized. In addition, highly viscous phosphatidylcholine gels can be obtained in organic solvents upon solubilizing a critical amount of water. Generally, phospholipid-based reverse micelles can be regarded as thermodynamically stable models for inverted micellar lipid structures possibly occurring in biological membranes.

Infrared Characterization of Different Water Types inside Reverse Micelles

The three Gaussian components of the IR stretching band of OH for reverse micelles of egg yolk L-α-phosphatidylcholine in benzene have been characterized in frequency, bandwidth, and extinction coefficient. The different types of water identified inside the micelles correspond to hydration layers around the phospholipid polar heads, the first containing up to ∼11 water molecules per polar head, the second with a maximum of 10–12 water molecules per polar head.

A spectroscopic study of poly(dA–dT)·poly(dA–dT) in microemulsions

The behaviour of high molecular weight [poly(dA–dT)·poly(dA–dT)] (polyAT) [(0.9–1.9)×106 Da] in the quaternary water-in-oil microemulsion CTAB–n-pentanol–n-hexane–water has been studied as a function of temperature and NaCl concentration by means of UV and circular dichroism (CD) spectroscopy. Solubilization in the microemulsion stabilises the polymer double helix and no helix-to-coil thermal transition has been observed. At increasing NaCl concentration, the CD spectrum of polyAT exhibits dramatic changes and for salt concentrations 7.5×10−1 M the spectral pattern suggests the formation of aggregates of the ψ(−) type. Condensation is induced by the peculiar characteristics of the aqueous core of the microemulsion. The condensates seem to destabilise the microemulsive system and, with time, separate from it.

Different factors affecting polyAT conformation in microemulsions: effects of variable P0 and KCl concentration

The behaviour of the synthetic polynucleotide polyAT entrapped in the inner core of a quaternary cationic microemulsion has been investigated by means of CD and UV spectroscopies in the presence of KCl (variable temperature at constant W0 = [H2O]/[CTAB] and P0 = [cosurfactant]/[CTAB]) and NaCl (variable P0 at constant W0 and temperature). While the presence of KCl gives rise to effects which are not substantially different from those already reported for NaCl (absence of thermal melting and formation of compact ψ(−) structures), the peculiar results obtained as a consequence of the progressive addition of cosurfactant (denaturation of the polymer at low [NaCl] and W0; transition from B- to A-form at intermediate [NaCl], formation of ψ(−) aggregates at high [NaCl]) can be attributed to the characteristics of the microemulsive system (restricted volume), of the micellar water (low dielectric constant and reduced activity) and of the interface (progressively higher cosurfactant concentration).

Acid Titrations of poly(dG-dC).poly(dG-dC) in Aqueous Solution and in a w/o Microemulsion

Abstract The model polynucleotide poly(dG-dC).poly(dG-dC) (polyGC) was titrated with a strong acid (HCl) in aqueous unbuffered solutions and in the quaternary w/o microemulsion CTAB/n-pentanol/n-hexane/water. The titrations, performed at several concentrations of NaCl in the range 0.005 to 0.600 M, were followed by recording the modifications of the electronic absorption and of the CD spectra (210≤ λ ≤350 nm) upon addition of the acid. In solution, the polynucleotide undergoes two acid-induced transitions, neither of which corresponds to denaturation of the duplex to single coil. The first transition leads to the Hoogsteen type synG.C+ duplex, while the second leads to the C+.C duplex. The initial B- form of polyGC was recovered by back-titration with NaOH. The apparent pKa values were obtained for both steps of the titration, at all salt concentrations. A reasonably linear dependence of pKa(1) and pKa(2) from p[NaCl] was obtained, with both pKa values decreasing with increasing ionic strength. In microemulsion, at salt concentrations ≤ 0.300 M, an acid-induced transition was observed, matching the first conformational transition recorded also in solution. However, further addition of acid led to denaturation of the protonated duplex. Renaturation of polyGC was obtained by back-titration with NaOH. At salt concentrations > 0.300 M, polyGC is present as a mixture of B-form and ψ(-) aggregates, that slowly separate from the microemulsion. The acid titration induces at first a conformational transition similar to the one observed at low salt or in solution, then denaturation occurs, which is however preceded by the appearance of a transient conformation, that has been tentatively classified as a left-handed Z double helix.

PolyAT chemical denaturation in w/o microemulsion

CD and UV spectroscopies have been used to investigate the effects caused by the addition of either strong acid-or base-containing microemulsions on the behaviour of the synthetic polynucleotide polyAT entrapped in the aqueous core of a cationic quaternary water-in-oil microemulsion (μE). The titrations were performed in the presence of variable concentrations of NaCl, in the range 0.00 to 0.60 M. In both cases, the primary effect was the reversible transition from B-double helix to random coil of the guest polynucleotide. However, in the microemulsive medium, the number of moles of protons (RH) and hydroxide ions (ROH) per mole of titrable sites are independent of the salt concentration but larger than 0.5, the value predicted on the basis of the stoicheiometry of the protonation-deprotonation processes. This result is in contrast with that obtained in aqueous solution (higher RH and ROH values and strongly dependent on NaCl concentration) and is explained with the presence of the cationic micellar wall (CTAB polar heads) acting as a ionic strength buffer.

Towards NMR Spectroscopy In Vivo: II. Relationship between NMR Parameters and Histology

The future of NMR as an in vivo diagnostic tool, both for imaging techniques and for spectroscopic determinations, requires as a preliminary step a precise tissue characterization. In practice, sound relationships should be established between the histology of a tissue and the relevant NMR parameters.

Towards NMR Spectroscopy In Vivo : The Use of Models

In recent years the use of NMR for clinical purposes has been increasing. Magnetic resonance imaging has great diagnostic potential, despite the many problems and drawbacks involved in its practice (1). More controversial is the possibility to exploit NMR spectroscopy as a diagnostic tool in clinical practice. Its applications seem to be restricted to the study of phosphorus containing metabolites (2), in particular to muscle energetics and to the viability of organs. However, some very recent research, based on 1H (3) and 13C (4,5) NMR, opens new fields in the applications of in vivo NMR to clinical problems.