Gemma Montalvo

Effect of fatty acids on self-assembly of soybean lecithin systems.

With the increasing interest in natural formulations for drug administration and functional foods, it is desirable a good knowledge of the phase behavior of lecithin/fatty acid formulations. Phase structure and properties of ternary lecithin/fatty acids/water systems are studied at 37°C, making emphasis in regions with relatively low water and fatty acid content. The effect of fatty acid saturation degree on the phase microstructure is studied by comparing a fully saturated (palmitic acid, C16:0), monounsaturated (oleic acid, C18:1), and diunsaturated (linoleic acid, C18:2) fatty acids. Phase determinations are based on a combination of polarized light microscopy and small-angle X-ray scattering measurements. Interestingly, unsaturated (oleic acid and linoleic acid) fatty acid destabilizes the lamellar bilayer. Slight differences are observed between the phase diagrams produced by the unsaturated ones: small lamellar, medium cubic and large hexagonal regions. A narrow isotropic fluid region also appears on the lecithin-fatty acid axis, up to 8wt% water. In contrast, a marked difference in phase microsctructure was observed between unsaturated and saturated systems in which the cubic and isotropic fluid phases are not formed. These differences are, probably, a consequence of the high Krafft point of the C16 saturated chains that imply rather rigid chains. However, unsaturated fatty acids result in more flexible tails. The frequent presence of, at least, one unsaturated chain in phospholipids makes it very likely a better mixing situation than in the case of more rigid chains. This swelling potential favors the formation of reverse hexagonal, cubic, and micellar phases. Both unsaturated fatty acid systems evolve by aging, with a reduction of the extension of reverse hexagonal phase and migration of the cubic phase to lower fatty acid and water contents. The kinetic stability of the systems seems to be controlled by the unsaturation of fatty acids.

Structure and phase equilibria of the soybean lecithin/PEG 40 monostearate/water system

PEG stearates are extensively used as emulsifiers in many lipid-based formulations. However, the scheme of the principles of the lipid-surfactant polymer interactions are still poorly understood and need more studies. A new phase diagram of a lecithin/PEG 40 monostearate/water system at 30 °C is reported. First, we have characterized the binary PEG 40 monostearate/water system by the determination of the critical micelle concentration value and the viscous properties. Then, the ternary phase behavior and the influence of phase structure on their macroscopic properties are studied by a combination of different techniques, namely, optical microscopy, small-angle X-ray scattering, differential scanning calorimetry, and rheology. The phase behavior is complex, and some samples evolve even at long times. The single monophasic regions correspond to micellar, swollen lamellar, and lamellar gel phases. The existence of extended areas of phase coexistence (hexagonal, cubic, and lamellar liquid crystalline phases) may be a consequence of the low miscibility of S40P in the lecithin bilayer as well as of the segregation of the phospholipid polydisperse hydrophobic chains. The presence of the PEG 40 monostearate has less effect in the transformation to the cubic phase for lecithin than that found in other systems with simple glycerol-based lipids.

Analysis of street cocaine samples in nasal fluid by Raman spectroscopy.

The principal objective of this work was to demonstrate the capability of Raman spectroscopy to detect small amounts of cocaine in nasal fluid, and to identify the main drug and the most widely used cutting agents. Initially, standard samples were analysed and sampling conditions were studied by comparing different swabs used for the sample collection. Once the most appropriate swab was selected, which permitted a relatively simple detection of the standard cocaine hydrochloride, qualitative analyses of real samples were carried out. Three street cocaine samples were analysed, and the presence of cutting substances was highlighted by the appearance of different bands not corresponding to the ones of the standard cocaine. To identify the substances present in each sample, the spectra of the street cocaine samples were collected and compared with a digital library created on purpose with the spectra of the most common cutting agents. In this case, correlation coefficients permitted to recognize the most important substances presumably present in the samples, and gave an estimation of the purity of the cocaine. However, when nasal fluid was present, its strong signal could overlap or interfere with the smaller signal of the cutting substances, hindering their identification.

Forensic discrimination of inkjet-printed lines by Raman spectroscopy and surface-enhanced Raman spectroscopy

Tracing a printed document back to its device is an important issue in forensic examinations of questioned documents. Raman spectroscopy is a resourceful technique for document analysis since it provides molecular information of many inks’ components, the type of paper has no effect on the results, and it is a non-destructive technique. Research studies utilise Raman instruments that have a greater potential than the ones used by many law enforcement institutions, which are adapted for routine casework. The potential and limitations were investigated of one Raman instrument, which was especially designed for document casework with regards to the complex discrimination of samples printed by different inkjet printers from the same manufacturer. The results showed low signals for most of the samples, especially black lines, due to fluorescence. To eliminate this, surface-enhanced Raman spectroscopy (SERS) was performed. The surface treatment method with a silver colloid was studied and optimised to be a less invasive method. SERS spectra resulted in more intense and well-shaped bands but the same profile, typical of copper phthalocyanine derivatives, overwhelmed the entire spectra for blue and black ink lines. Notwithstanding, closer examination revealed smaller spectral differences, which allowed discrimination of the samples into two groups: samples printed by the Deskjet models and samples printed by professional models such as Photosmart, Business and Officejet.

Structure of aggregates in diluted aqueous octyl glucoside/tetraethylene glycol monododecyl ether mixtures with different alkanols.

A systematic study of the diluted lamellar phases of the OG/C(12)E(4) system with different alkanols has been carried out by small-angle X-ray scattering (SAXS). The measurements have been made as a function of both the concentration and the alcohol type. Several different form factor models have been used to estimate the differences in bilayer topology induced by the presence of alcohol. For the infinite lamellae form factor (with a high-low-high electronic density profile across the membrane), there is a good fitting of samples with a X(OG) = 0.1 ratios. Only the free parameters correspond to the pseudomolecule composition and hydration number, which resulted in two water molecules per ethylene oxide group in the polar head irrespective of the alkanol chain length and concentration. However, samples with higher OG content can be quite well fitted by a core-shell disk model. For the samples with higher OG content, we find the participation of OG in the disks to be important. From the line-shape analysis of SAXS data, the half-thickness of the hydrophobic layer and the thickness of the hydrophilic layer have also been obtained. The results suggest significant mixing of the surfactant acyl chains corresponding to both sides of the lamellae and the transition from vesicles to open bilayer fragments without macroscopic phase separation.

Ultraviolet resonance Raman spectroscopy for the detection of cocaine in oral fluid

Detecting and quantifying cocaine in oral fluid is of significant importance for practical forensics. Up to date, mainly destructive methods or biochemical tests have been used, while spectroscopic methods were only applied to pretreated samples. In this work, the possibility of using resonance Raman spectroscopy to detect cocaine in oral fluid without pretreating samples was tested. It was found that ultraviolet resonance Raman spectroscopy with 239-nm excitation allows for the detection of cocaine in oral fluid at 10μg/mL level. Further method development will be needed for reaching the practically useful levels of cocaine detection.

Raman spectral signatures for the differentiation of benzodiazepine drugs

The identification of benzodiazepine drugs is important in the forensics field because they are used in drug-facilitated crimes. Raman spectroscopy has been proven as a non-invasive, fast and reliable technique highly promising for the analysis of drug products. Until recently, attention has been paid to active ingredients, but the spectral drug product signature has rarely been used in spite of having potential valuable information. In this work, confocal Raman microscopy was used to obtain the spectral signature of the most widely used benzodiazepine products. Firstly, the study aimed at determining appropriate Raman mapping spectra to obtain each benzodiazepine signature with low sampling error. Then, PCA scores and loadings showed that the variability, measured on the variance, among batches of the same benzodiazepine drug was similar to the variability of the spectral signature of the same tablet (or capsule content) and the same batch, mainly attributed to the heterogeneity of such drug products. Interestingly, differentiation among doses of the same active ingredient (AI), benzodiazepine drugs with different AIs manufactured by the same pharmaceutical company, and drugs with the same AI but manufactured by different companies were demonstrated. It is remarkable that for low doses, the active ingredient is almost absent of the spectral signature, but the differentiation is mainly achieved by excipients. As a consequence, the spectral signature obtained by confocal Raman microscopy can be used for discriminating among these benzodiazepine drugs without requiring a clearly identifiable band related to the active ingredient in the corresponding Raman spectra.

Effect of Meso vs Macro Size of Hierarchical Porous Silica on the Adsorption and Activity of Immobilized β-Galactosidase

β-Galactosidase (β-Gal) is one of the most important enzymes used in milk processing for improving their nutritional quality and digestibility. Herein, β-Gal has been entrapped into a meso-macroporous material (average pore size 9 and 200 nm, respectively) prepared by a sol-gel method from a silica precursor and a dispersion of solid lipid nanoparticles in a micelle phase. The physisorption of the enzyme depends on the concentration of the feed solution and on the pore size of the support. The enzyme is preferentially adsorbed either in mesopores or in macropores, depending on its initial concentration. Moreover, this selective adsorption, arising from the oligomeric complexation of the enzyme (monomer/dimer/tetramer), has an effect on the catalytic activity of the material. Indeed, the enzyme encapsulated in macropores is more active than the enzyme immobilized in mesopores. Designed materials containing β-Gal are of particular interest for food applications and potentially extended to bioconversion, bioremediation, or biosensing when coupling the designed support with other enzymes.

Monitoring of the stability of cocaine and some metabolites in water and oral fluid by a newly developed CE method

A new CE method was here developed, in order to study the stability of cocaine and some of its metabolites in water and in oral fluid. At first, standard mixtures of cocaine (COC), benzoylecgonine (BE) and cocaethylene (COET) in water were used to study the optimal CE parameters to separate the three compounds. Voltage, sample temperature and pH were investigated, and 25 kV, 25°C and a pH of 4.7 were selected to achieve the best separation. The stability of the three compounds in water and oral fluid was then monitored by applying the previously developed method. Three different storage temperatures (8, 25 and 37°C) were selected and analyses during a week were performed. A decrease of COC and COET peak areas and an increase of BE peak area were observed over time at 25 and 37°C. In addition, in oral fluid, the presence of enzymes and other proteins, and the differences in the molecular structures between COC and COET, caused a stronger degradation of the first compound. Instead, when samples were stored at a low temperature (8°C), the peak areas of the compounds did not vary. Thus, the use of this storage temperature is recommended, above all when sample must be analyzed after a relatively long time.

Gelatin/Maltodextrin Water-in-Water (W/W) Emulsions for the Preparation of Cross-Linked Enzyme-Loaded Microgels

Cross-linked gelatin microgels were formed in gelatin-in-maltodextrin water-in-water (W/W) emulsions and evaluated as carriers of the enzyme β-galactosidase (β-Gal). The phase behavior of aqueous gelatin/maltodextrin mixtures was studied in detail, focusing on the multiphase region of the phase diagram that is constituted by three equilibrium phases: two immiscible aqueous phases plus one solid phase. The solid phase was analyzed by Raman spectroscopy, and water-in-water emulsions were formed within the multiphase region. Gelation of the dispersed gelatin droplets was induced by cooling and cross-linking with genipin, which is a natural cross-linking reagent of low toxicity, leading to the formation of gelatin microgel particles. These microgels were studied as delivery vehicles for the enzyme lactase, used as a model active component. Various incorporation methods of the enzyme were tested, to achieve highest encapsulation yield and activity recovery. Microgel particles, loaded with the enzyme, can be freeze-dried, and the enzyme remained active after a complete cycle of freeze-drying and rehydration. The stability of the enzyme at 37 °C under gastric and neutral pH conditions was tested and led to the conclusion that the cross-linked microgels could be suitable for use in food-industry, where β-Gal carriers are of interest for hydrolyzing lactose in milk products.