Spray-drying of protein/polysaccharide complexes: dissociation of the effects of shearing and heating steps Jian Wang, Faydi Maoulida, Chedia Ben Amara, Emilie Dumas, Adem Gharsallaoui Univ Lyon 1, ISARA Lyon, Laboratoire BioDyMIA, Equipe Mixte d’Accueil, n°3733, IUT Lyon 1, technopole Alimentec, rue Henri de Boissieu, F-01000 Bourg en Bresse, France adem.gharsallaoui@univ-lyon1.fr EFW 2018: The 10th International Conference on Water in Food, 19–21 September 2018, Prague (Czech Republic)

Introduction Experimental design Effect of atomization on the properties of caseinate/pectin complexes Effect of heat treatment on the properties of caseinate/pectin complexes Combined effect of atomization and heat treatment of complexes Conclusion Plan

1. Introduction : Spray-drying process Air Peristaltic pump Nozzle Drying air Atomization Heating Spray-drying : efficient and economical process which was widely used in food industry; The spray-drying process can be divided into two steps: atomization and heating. Atomization step : pulverization of the liquid into small droplets; Heating step : the droplets come into contact with heat air  evaporation of water.

Polysaccharides in solution Coacervates / Precipitation 1. Introduction : About complex coacervation (in aqueous medium) Proteins in solution Polysaccharides in solution + Attractive interactions Repulsive interactions Soluble complexes Coacervates / Precipitation C Incompatibility Co-solubility C pH, I Spray-Drying PROCESS ? 4

Aims of the present work 1. Introduction Aims of the present work Dissociating mechanical and thermal steps during Spray-drying. Studying separately the effect of atomization on complexes properties. Studying separately the effect of heat treatment on complexes properties. Evaluation of the effect of atomization followed by heating on complexes structure.

2. Experimental Design According to the effect of pH on the zeta-potential of caseinate and pectin, at pH 3, caseinate is positively charged, and pectin is negatively charged, complex coacervation can be easily formed at pH 3 by electrostatic force. Sodium caseinate and Low methoxyl pectin are chosen as the protein/polysaccharide system; Zeta-potential  electrostatic complexes are formed at pH 3 (oppositely charged biopolymers). 6

2. Experimental Design Heat treatment* Heat treatment* Pectin Atomization Imidazole-acetate buffer (5 mM) at pH 3 Pectin Caseinate Final pectin concentration: 0.5 g/L, 2.0 g/L and 6.0 g/L Constant Caseinate concentration: 5.0 g/L Caseinate/Pectin complexes Caseinate/Pectin complexes after atomization Turbidity; Zeta-potential; Particle size distribution; Surface hydrophobicity. Complexes after atomization followed by heat treatment Heat treatment* Caseinate/Pectin complexes after heat treatment Heat treatment* *Heat treatment: water bath at 80°C for: 1 min (T1), 2 min (T2), 3 min (T3) and 4 min (T4). Control : Unheated complexes (T0).

3. Effect of atomization on the properties of caseinate/pectin complexes Imidazole-acetate buffer (5 mM) at pH 3 Pectin Caseinate Final pectin concentration: 0.5 g/L, 2.0 g/L and 6.0 g/L Constant Caseinate concentration: 5.0 g/L Caseinate/Pectin complexes Caseinate/Pectin complexes after atomization Turbidity; Zeta-potential; Particle size distribution; Surface hydrophobicity. Complexes after atomization followed by heat treatment Heat treatment* Caseinate/Pectin complexes after heat treatment Heat treatment* *Heat treatment: water bath at 80°C for: 1 min (T1), 2 min (T2), 3 min (T3) and 4 min (T4). Control : Unheated complexes (T0).

3. Effect of atomization on the properties of caseinate/pectin complexes Before atomization : the turbidity of complexes increased with increasing pectin concentration while the average size reached a peak  at 6.0 g/L of pectin, formation of small complexes but more numerous; After atomization : partial dissociation of complexes due to shear forces and electrostatic repulsions between the pectin chains surrounding the complexes; Complexes formed at high pectin concentrations are more sensitive to atomization.

3. Effect of atomization on the properties of caseinate/pectin complexes Increasing pectin concentration: the electrostatic charges changed from positive to negative  adsorption of pectin and formation of electrostatic complexes; Atomization : the dissociation of the complexes after atomization is due to: - Shear forces during their passage in the nozzle; - An increase in their overall charge favoring electrostatic repulsions.

3. Effect of atomization on the properties of caseinate/pectin complexes Before atomization: slight decrease of the protein hydrophobicity only at high pectin concentration (6.0 g/L)  the hydrophobic sites of caseinate would be masked by the pectin chains; After atomization: a significant increase of S0 was observed (particularly at 2.0 g/L and 6.0 g/L pectin):  Hypothesis : molecular rearrangements and unfolding of the structure of the protein  exposure of hydrophobic sites.

4. Effect of heat treatment on the properties of caseinate/pectin complexes Atomization Imidazole-acetate buffer (5 mM) at pH 3 Pectin Caseinate Final pectin concentration: 0.5 g/L, 2.0 g/L and 6.0 g/L Constant Caseinate concentration: 5.0 g/L Caseinate/Pectin complexes Caseinate/Pectin complexes after atomization Turbidity; Zeta-potential; Particle size distribution; Surface hydrophobicity. Complexes after atomization followed by heat treatment Heat treatment* Caseinate/Pectin complexes after heat treatment Heat treatment* *Heat treatment: water bath at 80°C for: 1 min (T1), 2 min (T2), 3 min (T3) and 4 min (T4). Control : Unheated complexes (T0).

4. Effect of heat treatment on the properties of caseinate/pectin complexes With increasing heat treatment duration from 0 to 4 min: 0.5 g/L of pectin: the turbidity increased with the heating time and there was no significant effect on the size of complexes  Assumption : aggregation of free caseinate molecules (protein denaturation); 2.0 g/L: the turbidity remained stable as a function of the heating time but the size of complexes increased  aggregation of complexes due to heat treatment; 6.0 g/L: the turbidity increased and the size of the complexes decreased  dissociation of the aggregates. Schéma

4. Effect of heat treatment on the properties of caseinate/pectin complexes With increasing heat treatment duration from 0 to 4 min: 0.5 g/L: Slight decrease in zeta-potential and increase in surface hydrophobicity: the structure of complexes changed (exposure of anionic groups of pectin and hydrophobic sites in caseinate molecules (denaturation)); 2.0 g/L: change of the electrostatic charge from positive to negative and a slight increase in hydrophobicity : the molecular rearrangement allowed to “protect” the protein from heat denaturation; 6.0 g/L: S0 is almost constant as a function of heating time: protective effect of pectin at high pectin concentrations. For zeta potential, 2 g/L : it’s clearly different, For the other one, is it different (stat’?)? For the second sentence, Some (;), some (:) : I find that this is complicated

5. Combined effect of atomization and heat treatment of complexes Imidazole-acetate buffer (5 mM) at pH 3 Pectin Caseinate Final pectin concentration: 0.5 g/L, 2.0 g/L and 6.0 g/L Constant Caseinate concentration: 5.0 g/L Caseinate/Pectin complexes Caseinate/Pectin complexes after atomization Turbidity; Zeta-potential; Particle size distribution; Surface hydrophobicity. Complexes after atomization followed by heat treatment Heat treatment* Caseinate/Pectin complexes after heat treatment Heat treatment* *Heat treatment: water bath at 80°C for: 1 min (T1), 2 min (T2), 3 min (T3) and 4 min (T4). Control: Unheated complexes (T0).

5. Combined effect of atomization and heat treatment of complexes With increasing heat treatment duration from 0 to 4 min: 0.5 g/L: slight increase in turbidity and constant size of the complexes  increase in the number of complexes due to the aggregation of free or partially complexed caseinate molecules; 2.0 g/L: decrease in turbidity and significant increase in size  association between small complexes accentuated by electrostatic charge neutralization; 6.0 g/L: turbidity increased while the size did not change  thermal dissociation of aggregates (already deconstructed by shearing) and formation of new complexes with free pectin chains.

Heat treatment after atomization 6. Conclusion 0.5 g/L 2.0 g/L 6.0 g/L 0.5 g/L 2.0 g/L 6.0 g/L At low concentration of pectin, heat treatment causes the aggregation of excess caseinate; + + + + + + + + + + Heat treatment + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + At intermediate concentration of pectin, heat treatment intensifies aggregation of complexes; At high concentration of pectin, heat treatment favors the dissociation of the aggregates. + + + + + + + + + + + + + + + + + + + + + + + + Atomization Atomization causes dissociation of aggregates into smaller size complexes at all pectin concentrations. Complexes after atomization are more sensitive to heat treatment; A high concentration of pectin can protect the complexes against heat aggregation. + + + + + + + + + + + + + + + + + + + + + + + + + + Heat treatment after atomization + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + + 0.5 g/L 2.0 g/L 6.0 g/L 0.5 g/L 2.0 g/L 6.0 g/L

6. Conclusion Future works : To better characterize the nature of the protein-polysaccharide binding, other complementary techniques like FTIR (Fourier transform infrared spectroscopy), NMR (nuclear magnetic resonance) and CD (circular dichroism) can be used for further study; Apply caseinate/pectin system to encapsulate some bio-active molecules like essential oils by Spray-drying.

THANK FOR YOUR ATTENTION Bourse chinoise à ajouter?