Gustavo González-Gaitano

The Aggregation of Cyclodextrins as Studied by Photon Correlation Spectroscopy

Photon correlation spectroscopy has been used to study theaggregation processes of natural and some modified cyclodextrins (CDs) in diluted aqueoussolutions. α-, β-, and γ-CD form large, polydisperse aggregatesin water, although the aggregation capability is different depending on the macrocycle considered. γ-CD solutions filtered through 0.2~μm give a single-modaldistribution of aggregates of 224 nm in size. The monomeric γ-CD can be isolated by filtering through 0.1 μm. α-CD displays a bimodaldistribution (monomer + aggregates) with both pore sizes. At the concentrations studied (0.012 M) the contribution in mass of the aggregates with both CDs isnegligible. β-CD is much more persistent in its aggregation, even after sievingits solutions through 0.02 μm filters, and time dependent. The aggregation displayed byCDs with partial substitution of the OH groups (Methyl-β-CD and Hydroxypropyl-β-CD) is much weaker, indicating the implication of the hydrophilic rims ofthe CDs in the process. High temperatures, addition of urea or electrolytes andionisation of the OH groups by raising the pH, prevent the aggregation.

Sorption models in cyclodextrin polymers: Langmuir, Freundlich, and a dual-mode approach

The classic Langmuir and Freundlich sorption models and a dual-mode approach have been tested to study the sorption of aromatic molecules onto beta-cyclodextrin polymers as well as onto analogous sucrose polymers, obtained using the same crosslinking agents (epichlorohydrin, succinyl chloride, toluene diisocyanate, and hexamethylene diisocyanate). The host-guest interaction of the sorbate within the cyclodextrin cavities corresponds to the hole-filling mechanism considered in the dual-mode approach, while the polymer crosslinking networks are capable of entrapping more sorbate molecules via partition. In some cases, when the sorption is governed by the inclusion within the cyclodextrin moieties, a simple Langmuir isotherm fits the data properly. The classic Freundlich equation is also appropriate when phenol is the sorbate because its interaction with beta-cyclodextrin is less specific than that of 1-naphthol.

Complementary multianalytical approach to study the distinctive structural features of the main humic fractions in solution: Gray humic acid, brown humic acid, and fulvic acid

Previous studies have indicated that the main fractions of humic substances (HS), gray humic acid (GHA), brown humic acid (BHA), and fulvic acid (FA), present different molecular patterns in water solution that are probably associated with specific structural features. However, the techniques used in these previous studies did not permit clarification of the principal qualitative characteristics of these structures. To study more in depth this subject several GHA, BHA, and FA have been analyzed through the complementary use of UV-visible and FTIR spectroscopy, (13)C NMR, thermogravimetry, and pyrolysis GC-MS. The results indicate that the studied humic fractions have different and distinctive structural features. Thus, large and nonpolar structural units (paraffins, olefins, terpenes) and aliphatic structures seem to accumulate in the gray fraction, whereas the smallest and more polar (furfural, phenols) and simpler structural units (sugar- and amino acid-related structures) are present in the fulvic one. BHA has a higher content in polycyclic aromatic moieties, S-containing compounds and aromatic structures, thus suggesting the presence of more condensed aromatic rings. Likewise, differences in both the presence of polar groups and the apparent molecular size explain the pattern of solubility as a function of pH and ionic strength (I) that defines each HS fraction. These results also indicate that the structural differences among the HS fractions are not only quantitative (the presence of the same type of structures differing in size and the concentration of functional groups) but also qualitative, because each fraction presented different and distinctive structural domains. These structural domains explain the molecular patterns associated with each HS fraction. Thus, the presence of smaller and more O-functionalized structural units including aromatic domains in FA explain their tendency to form molecular aggregates (hydrogen bridges, metal bridges, and hydrophobic interactions) in solution. This fact could also explain the presence of molecular aggregates in BHA, although to a lesser extent than in FA. Finally, the dominant aliphatic and less functionalized character of GHA may justify its lower tendency to form aggregates in solution at neutral and alkaline pH. Likewise, the results also indicate that the different structural domains associated with these fractions may be the consequence of diverse biosynthetic pathways involving different precursors.

Natural cyclodextrins as efficient boosters of the chemiluminescence of luminol and isoluminol: exploration of potential applications.

The chemiluminescent oxidation of luminol (LUM) and isoluminol (ISOL) is notably enhanced, both in intensity and duration, in the presence of natural cyclodextrins (alpha-, beta-, gamma-CD). The experiments have considered some of the most widespread applications of these compounds: the determination of metal cations and the revealing of bloodstains by oxidation with hydrogen peroxide in alkaline solution in the presence of Co(II), Fe(III), human hemoglobin, and blood, in order to explore potential applications. The largest enhancement in the emitted intensity occurs for the reaction of LUM with Co(II) in the presence of beta-CD. The use of the more soluble gamma-CD permits to expand the range of concentration and obtain more intense emission, although soluble derivatives of the beta-CD (methyl, hydroxypropyl-beta-CD, and a soluble cross-linked epichlorhydrin polymer) do not improve the chemiluminescence (CL) yield. In the case of hemoglobin and diluted human blood, the CDs aid in producing more light but only at high concentration of CDs, with a more lasting luminescence, up to three times longer. The changes in CL when glucose is used instead, much lower than with any of the CDs, imply that the cyclic structure of these oligosaccharides plays a key factor in the boosting of the emission. The results are explained in terms of the binding between the luminescent intermediate of the reaction, 3-aminophthalate (3-AP) and the CD, rather than to the luminescent reactant itself. The association constants obtained by steady-state fluorescence by assuming 1:1 stoichiometries reveal that the most stable association occurs between beta-CD and the intermediate, in accordance with the trend in the chemiluminescence. The topology of the complex deduced via ROESY experiments confirms a shallow inclusion of the double-charged intermediate by the primary rim of the CD, which accounts for the low stability of the complexes.

Fluorescence quenching investigation of the complexes of dibenzofuran with natural cyclodextrins

The steady-state fluorescence of dibenzofuran (DBF) in aqueous solution is quenched in the presence of natural cyclodextrins (CDs), α, β, and γ-CD. The decrease in the fluorescence is static in nature for all the macrocycles and follows as a result of the formation of inclusion complexes of dominant 1:1 stoichiometry. With γ-CD, excimer emission is detected at high CD concentrations due to the formation of a complex of 2:2 stoichiometry, DBF2:γ-CD2. The binding constants have been determined from the fluorescence intensities through the analysis of the F0/F vs. [CD] plots, resulting in a stronger association for β-CD. The thermodynamic parameters of the binding, enthalpy and entropy, have been calculated from the temperature dependence of the formation constants. The different trend displayed by α-CD, in both the absorption and fluorescence spectra, seems to be related to a shallow inclusion of DBF, stabilized by hydrogen bonding between the oxygen atom of the furan ring and the hydroxyl groups of CD.

Selective tuning of the self-assembly and gelation of a hydrophilic poloxamine by cyclodextrins

Complexes formed between cyclodextrins (CDs) and polymers - pseudopolyrotaxanes (PPRs) - are the starting point of a multitude of supramolecular structures, which are proposed for a wide range of biomedical and technological applications. In this work, we investigate the complexation of a range of cyclodextrins with Tetronic T1307, a four-arm block copolymer of poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) with a pH-responsive central ethylene diamine spacer, and its impact on micellization and the sol-gel transition. At low concentrations, small-angle neutron scattering (SANS) combined with dynamic light scattering (DLS) measurements show the presence of spherical micelles with a highly hydrated shell and a dehydrated core. Increasing the temperature leads to more compact micelles and larger aggregation numbers, whereas acidic conditions induce a shrinking of the micelles, with fewer unimers per micelle and a more hydrated corona. At high concentrations, T1307 undergoes a sol-gel transition, which is suppressed at pH below the pKa,1 (4.6). SANS data analysis reveals that the gels result from a random packing of the micelles, which have an increasing aggregation number and increasingly dehydrated shell and hydrated core with the temperature. Native CDs (α, β, γ-CD) can complex T1307, resulting in the precipitation of a PPR. Instead, modified CDs compete with micellization to an extent that is critically dependent on the nature of the substitution. (1)H and ROESY NMR combined with SANS demonstrate that dimethylated β-CD can thread onto the polymer, preferentially binding to the PO units, thus hindering self-aggregation by solubilizing the hydrophobic block. The various CDs are able to modulate the onset of gelation and the extent of the gel phase, and the effect correlates with the ability of the CDs to disrupt the micelles, with the exception of a sulfated sodium salt of β-CD, which, while not affecting the CMT, is able to fully suppress the gel phase.

Enhancement of the chemiluminescence of two isoluminol derivatives by nanoencapsulation with natural cyclodextrins.

The chemiluminescence (CL) yield of two isoluminol derivatives, N-(4-aminobutyl)-N-ethylisoluminol (ABEI) and N-(6-aminohexyl)-N-ethylisoluminol (AHEI), is remarkably increased in the presence of natural cyclodextrins (CDs). The most notable effect is produced by the addition of gamma-CD that produces enhancements up to 15-fold in the light emission of both compounds. Although proton nuclear magnetic resonance ((1)H NMR) measurements prove the encapsulation of these luminescent reagents in the CDs, with more stable associations by decreasing the width of the CD cavity, the improvement in the light emission of ABEI and AHEI is mainly due to the topology of the complexes. Evaluation of the rotating frame Overhauser effect spectroscopy (ROESY) spectra cross-peaks combined with semirigid-docking simulations has been used to gather information about the spatial conformation of the guest molecules into the CDs. These calculations have shown that a deeper inclusion in the CD cavity of the heterocyclic moiety of the luminescent molecules is directly related with a higher enhancement in the CL. The augment of the CL by natural CDs is of interest for increasing the detection limit in biochemical assays or liquid chromatography, for example, in which the CL of these compounds serves to quantify other molecular species that may take part, direct or indirectly, in the luminescent reaction.

Optimization of the entrapment of bacterial cell envelope extracts into microparticles for vaccine delivery

The encapsulation of a Brucella ovis extract (HS) in microparticles has been proved effective against experimental infections in mice. This work describes different strategies to increase the HS loading and prepare large batches as necessary to test this vaccine in ovine. The mixture of HS with β-cyclodextrin was optimized in order to increase the HS loading in microparticles. On the other hand, TROMS (‘Total Recirculation One-Machine System’) led microparticles with a more homogeneous size than the laboratory or standard procedure. Moreover, the initial burst release of HS from the standard microparticles was higher than for the TROMS ones. In fact, standard microparticles displayed a higher amount of adsorbed HS. On the contrary, both preparative methods were found effective to preserve the integrity and anti-genicity of the loaded HS. In summary, β-CD can be used to increase the loading of large hydrophobic materials and TROMS is a valid large production of antigen-loaded microparticles.

Electronic circuit for sustaining the oscillations of a vibrating element: Application to density measurements

An electronic circuit has been designed to sustain the oscillations of a vibrating element. It has been applied to density measurements of fluids with a vibrating tube densimeter. The electronic part of the apparatus has been designed to avoid the presence of elements attached to the tube (such as wires or magnets), since the tube itself performs the excitation and the detection, making possible an easy manufacture. The mechanical part permits the best temperature control and a fast thermal response, since it can be totally immersed in a thermostat. The precision has been tested by calibration with pure substances and applied to density measurements of a colloidal solution (sodium decanoate in water). Relative measurements are obtained with a precision of better than 1×10−6 g cm −3, in the temperature range studied (between 293 and 313 K).

New methodology to assess the quantity and quality of humic substances in organic materials and commercial products for agriculture

PurposeThe traditional method to determine humic content (humic and fulvic acids) in commercial fertilizers, biostimulants, and organic materials is based on the oxidation of the organic carbon contained in the basic-soluble but acid-insoluble fraction (humic acids) and the basic-acid soluble fraction (fulvic acids) of their alkaline water extracts. This methodology, merely operational, makes it impossible to distinguish if the quantified carbon corresponds to substances with “humic” chemical nature or to non-humic organic matter but with similar solubility properties to those of humic matter. The aim of this work is to develop a new methodology that not only quantifies the humic content in commercial products (and raw materials) but also assesses the humic quality of the quantified organic matter.Materials and methodsTo this end, humic and fulvic (-like) fractions have been isolated/purified from several humic and non-humic materials and characterized by means of elemental analysis and UV-visible, fluorescence, and infrared spectroscopies, and these data have been used to perform a discriminant analysis (DA).Results and discussionThe model obtained from the DA is able to discriminate humic and fulvic fractions from apparently humic or fulvic ones and provides discriminant classification functions that have proven to successfully predict the “humic quality” of the fractions isolated from commercial products, after their elemental and spectroscopic characterization.ConclusionsTherefore, the combination of the fractionation, characterization, and evaluation by the DA is proposed as an effective methodology for quantifying and assessing the quality of the humic content claimed in the labels of commercial products.