Patricio R. Santagapita

Amaranth Milling Strategies and Fraction Characterization by FT-IR

Amaranth nutritional value has been widely recognized, but the required conditions for its processing cannot be adapted to traditional technologies. For the proposal of alternative strategies, the changes of several components should be understood. Enriched starch and lipid–protein fractions of amaranth flour upon different milling treatments were obtained and characterized by attenuated total reflectance–Fourier transform-infrared spectroscopy. Starch- and lipid–protein-enriched amaranth fractions were obtained by abrasive milling; amaranth starch was isolated by wet-milling procedure, and flour samples were obtained from planetary ball milling. Changes on starch, protein, and lipids relative contents, on starch crystallinity and on lipids and protein stability after milling and 6-month storage, were evaluated. The Fourier transform-infrared (FT-IR) spectroscopy peaks of the main grain components were identified in the middle-infrared range. By calculating the ratios between height intensities of selected specific peaks, several characteristics of the samples could be explained: increased protein content and lipid proportion of the enriched fraction; decrease of the starch crystallinity degree by abrasive milling and especially by ball milling due to starch amorphization during these processes; and lipids modification in milled and in 6-month aged samples. FT-IR analysis can be considered a rapid, nondestructive, solvent-free, sensitive, and useful tool to investigate starch, lipid, and protein modifications provoked by processing and storage as well as to determine, based on intensity ratio, the relative proportion of grain components within amaranth milling fractions. The abrasive milling associated to planetary ball milling to obtain modified different fractions is presented as an interesting strategy for the processing of amaranth grain.

Encapsulation of lactase in Ca(II)-alginate beads: Effect of stabilizers and drying methods

The purpose of the present work was to analyze the effect of trehalose, arabic and guar gums on the preservation of β-galactosidase activity in freeze-dried and vacuum dried Ca(II)-alginate beads. Freezing process was also studied as a first step of freeze-drying. Trehalose was critical for β-galactosidase conservation, and guar gum as a second excipient showed the highest conservation effect (close to 95%). Systems with Tg values ~40°C which were stables at ambient temperature were obtained, being trehalose the main responsible of the formation of an amorphous matrix. Vacuum dried beads showed smaller size (with Ferets diameter below 1.08±0.09mm), higher circularity (reaching 0.78±0.06) and large cracks in their surface than freeze-dried beads, which were more spongy and voluminous. Ice crystallization of the beads revealed that the crystallization of Ca(II)-alginate system follows the Avrami kinetics of nucleation and growth. Particularly, Ca(II)-alginate showed an Avrami index of 2.03±0.07, which means that crystal growing is bidimensional. Neither the addition of trehalose nor gums affected the dimension of the ice growing or its rate. These results open an opportunity in the development of new lactic products able to be consumed by lactose intolerance people.

Gums induced microstructure stability in Ca(II)-alginate beads containing lactase analyzed by SAXS

Previous works show that the addition of trehalose and gums in β-galactosidase (lactase) Ca(II)-alginate encapsulation systems improved its intrinsic stability against freezing and dehydration processes in the pristine state. However, there is no available information on the evolution in microstructure due to the constraints imposed by the operational conditions. The aim of this research is to study the time course of microstructural changes of Ca(II)-alginate matrices driven by the presence of trehalose, arabic and guar gums as excipients and to discuss how these changes influence the diffusional transport (assessed by LF-NMR) and the enzymatic activity of the encapsulated lactase. The structural modifications at different scales were assessed by SAXS. The incorporation of gums as second excipients induces a significant stabilization in the microstructure not only at the rod scale, but also in the characteristic size and density of alginate dimers (basic units of construction of rods) and the degree of interconnection of rods at a larger scale, improving the performance in terms of lactase activity.

Encapsulation of a free-solvent extract of lycopene in alginate-Ca(II) beads containing sugars and biopolymers

AbstractBackgroundThe purpose of the present study was to enhance the stability toward isomerization and control the release of an encapsulated free-solvent extract of lycopene, obtained from a nonconventional natural source, by means of alginate beads containing sugar (trehalose) and biopolymers (chitosan, low methoxyl pectin, and arabic gum).MethodsLycopene was extracted from freeze-dried pulp of pink grapefruit obtaining a free solvent extract. Lycopene encapsulation was conducted by a double procedure consisting of emulsification and ionotropic gelation in alginate-Ca(II) beads, modified by the addition of sugar and biopolymers. The influence of beads’ composition was studied on lycopene stability and release, as well as molecular mobility and diffusion in the beads.Results and ConclusionsThe addition of a second excipient (besides alginate) in the formulation should be carefully conducted, since stability during alginate-Ca(II) bead generation could be even compromised, leading to high lycopene losses. Beads containing trehalose and chitosan were the ones that best preserved the lycopene content and minimized isomerization changes. This could be related to the reduced molecular mobility and lower diffusion coefficient of this system. Lycopene release was severely affected by the composition of the beads, allowing to modulate its release depending on a desired application. Then, a good strategy to obtain high lycopene formulations ready to use or for their incorporation in a subsequent technological process (such as freeze-drying or extrusion) was reported in the present study. Graphical abstractStability and release of encapsulated lycopene in alginate-Ca(II) beads

Impact of supramolecular interactions of dextran-β-cyclodextrin polymers on invertase activity in freeze-dried systems.

β‐Cyclodextrin (β‐CD)‐grafted dextrans with spacer arms of different length were employed to evaluate the impact of supramolecular interactions on invertase activity. The modified dextrans were used as single additives or combined with trehalose in freeze‐dried formulations containing invertase. Enzyme activity conservation was analyzed after freeze‐drying and thermal treatment. The change of glass transition temperature (Tg) was also evaluated and related to effective interactions. Outstanding differences on enzyme stability were mainly related to the effect of the spacer arm length on polymer–enzyme interactions, since both the degree of substitution and the molecular weight were similar for the two polymers. This change of effective interactions was also manifested in the pronounced reduction of Tg values, and were related to the chemical modification of the backbone during oxidation, and to the attachment of the β‐CD units with spacer arms of different length on dextran.

Polyethylene glycol‐based low generation dendrimers functionalized with β‐cyclodextrin as cryo‐ and dehydro‐protectant of catalase formulations

Polyethylene glycol (PEG)‐based low generation dendrimers are analyzed as single excipient or combined with trehalose in relation to their structure and efficiency as enzyme stabilizers during freeze‐thawing, freeze‐drying, and thermal treatment. A novel functional dendrimer (DGo‐CD) based on the known PEGs ability as cryo‐protector and β‐CD as supramolecular stabilizing agent is presented. During freeze‐thawing, PEG and β‐CD failed to prevent catalase denaturation, while dendrimers, and especially DGo‐CD, offered the better protection to the enzyme. During freeze‐drying, trehalose was the best protective additive but DGo‐CD provided also an adequate catalase stability showing a synergistic behavior in comparison to the activities recovered employing PEG or β‐CD as unique additives. Although all the studied dendrimers improved the enzyme remaining activity during thermal treatment of freeze‐dried formulations, the presence of amorphous trehalose was critical to enhance enzyme stability. The crystallinity of the protective matrix, either of PEG derivatives or of trehalose, negatively affected catalase stability in the freeze‐dried systems. When humidified at 52% of relative humidity, the dendrimers delayed trehalose crystallization in the combined matrices, allowing extending the protection at those conditions in which normally trehalose fails. The results show how a relatively simple covalent combination of a polymer such as PEG with β‐CD could significantly affect the properties of the individual components. Also, the results provide further insights about the role played by polymer–enzyme supramolecular interactions (host–guest crosslink, hydrogen bonding, and hydrophobic interactions) on enzyme stability in dehydrated models, being the effect on the stabilization also influenced by the physical state of the matrix.

Impact of protective agents and drying methods on desiccation tolerance of Salix nigra L. seeds.

Willow seeds are classified as orthodox, but they show some recalcitrant characteristics, as they lose viability in a few weeks at room temperature. The aim of this work was to improve the desiccation tolerance of willow seeds (Salix nigra L.), as a model of sensitive materials to dehydration, through imbibition in solutions and later vacuum (VD) or freeze-drying (FD). Imbibition was conducted with 45% w/v trehalose or polyethylene glycol 400 -PEG- or water prior to dehydration treatments. Water- and especially trehalose-imbibed seeds subjected to VD showed better germination capability with respect to the freeze-dried ones. Water crystallization was mainly responsible for the great loss of capability germination observed in water- or trehalose-imbibed seeds subjected to FD. PEG behavior was better when seeds were FD instead of VD. DSC thermograms of seeds allowed to identify two thermal transitions corresponding to lipids melting and to proteins denaturation. This last transition reveals information about proteins state/functionality. Dehydration of control and PEG- or water-imbibed seeds affected proteins functionality leading to lower germinability. In the case of trehalose-imbibed seeds subjected to VD, proteins maintained their native state along dehydration, and the seeds showed a great germination capacity for all the water content range. Germinated seeds showed higher luminosity (L*), greenness (a*) and yellowness (b*) values than not-germinated seeds independently of the employed agent. Present work reveals that the presence of adequate protective agents as well the dehydration method were the main critical factors involved in willow seed desiccation tolerance.