Obtención enzimática, caracterización y propiedades prebióticas de oligosacáridos empleados en leches infantiles
Autor (es)
Rodríguez Colinas, BárbaraEntidad
UAM. Departamento de Biología MolecularFecha de edición
2013-11-14Materias
Niños - Alimentación - Tesis doctorales; Oligosacáridos - Tesis doctorales; Leche - Tratamiento - Tesis doctorales; Instituto de Catálisis y Petroleoquímica (Madrid); Biología y Biomedicina / BiologíaNota
Tesis Doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de lectura: 14-11-2013Esta obra está bajo una licencia de Creative Commons Reconocimiento-NoComercial-SinObraDerivada 4.0 Internacional.
Resumen
Prebiotics are selectively fermented ingredients that allow specific changes in composition and/or
activity in the gastrointestinal microflora that confer benefits upon host well-being and health. In this
field, the attention is mainly focused on fructo-oligosaccharides (FOS) and galacto-oligosaccharides
(GOS) due to their current use in many foods and their importance in human health.
Human milk oligosaccharides (HMOs) supply all the nutritional needs to the baby and they are
essential to the proper development and growth of the baby. Their components provide bioactive
compounds that confer protection against pathogens and diseases, modulate immune system, stimulate
desired bacteria like Bifidobacterium and Lactobacillus, influence in cognitive development and
establishment of the intestinal microbiota. Due to the complexity of these HMOs, it is a common practice
the supplementation of FOS/GOS in infant formulas as alternative sources of prebiotic ingredients to
provide similar bifidogenic effects to breast fed babies.
In this work several enzymes were studied for their use in prebiotic oligosaccharide production;
three β-galactosidases from K. lactis, B. circulans and A. oryzae for GOS synthesis and one β-
fructofuranoside from A. aculeatus for FOS production. The major products synthesized by B. circulans
β-galactosidase contained only β (1→4) bonds, whereas the enzyme from K. lactis and A. oryzae
displayed a clear tendency to form β (1→6) linkages. B. circulans and K. lactis β-galactosidases produce
nearly 45-50% (w/w) GOS, whereas the A. oryzae enzyme produced less than 30% (w/w) GOS.
Regarding to product distribution B. circulans enzyme synthesized the highest amount of trisaccharides
wheras K. lactis and A. oryzae would be a better choice to synthesize a product with content in di- and
trisaccharides. The β-fructofuranoside from A. aculeatus formed basically a mixture of 1-kestose, nystose
and 1F-fructosylnystose.
It is reported that the chemical structure of the obtained oligosaccharides (composition, number of
hexose units and types of linkages between them) may affect their fermentation pattern by probiotic
bacteria in the gut.
To verify this effect, fermentation selectivity of some purified carbohydrates, with different
polymerization degree and distribution, was estimated using in vitro fermentation systems inoculated
with human fecal slurries. Changes in bacterial populations were measured by fluorescence in situ
hybridization (FISH) and HPLC analysis of short-chain fatty acids (SCFA) produced by oligosaccharide
fermentation. All the tested GOS could be considered bifidogenic under the assayed conditions with
selective index (SI) slightly lower to the determined for a commercial GOS mixture.
A challenging objective of this work was to obtain milk with a low content of lactose and a
significant amount of GOS. GOS concentration obtained with K. lactis β-galactosidase using skim milk
was close to the HMOs content in breast milk. Pasteurization after controlled enzymatic treatment could
result in a product with a low lactose content and with the extra benefit of a significant presence of
prebiotic GOS. In order to scale-up the processes to synthesize prebiotic oligosaccharides, immobilization in
calcium alginate gel beads was successfully performed for both β-galactosidase from B. circulans and β-
fructofuranoside from A. aculeatus. The controlled dehydration of alginate beads resulted in novel
biocatalysts (DALGEEs, Dry ALGinate Entrapped Enzymes) with improved properties with respect to
the wet beads. DALGEEs displayed higher volumetric activity, reduced volume and better storage. The
operational stability of DALGEEs containing β-fructofuranoside in a fixed-bed reactor was extraordinary
with a nearly constant FOS composition during 700 h.
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