Milk proteins, sodium caseinate (CN) and whey protein isolate (WPI) are used in food industries as emulsifiers. The stability of an O/W emulsion is dependent on the electrostatic and steric repulsion provided by the interfacial proteins against droplet aggregation or lipid oxidation. Therefore, modifications of the surface charge or the interfacial conformation of protein emulsifiers are expected to enhance their emulsifying properties and emulsion stability. In this present work, sodium caseinate and whey protein isolate were modified by different chemical and enzymatic approaches. The modified proteins were characterized using multiple techniques, and the effect of these modifications on emulsifying properties of proteins and emulsion stability were investigated. Succinylation converts the positively charged amino groups into negatively charged carboxyl groups, lowering the isoelectric point (pI) of protein. The ethylene diamine (EDA) modification worked in the opposite way, leading to an increased pI. The extent of these two modifications was studied using SDS-PAGE and MALDI-TOF mass spectrometry. The pI of succinylated and EDA modified milk proteins was studied using zeta-potential measurement. As a result, the succinylation to full extent altered the pI of CN from 4.2 to 2.7, and the EDA modification shifted the pI of CN and WPI from 4.2 to 9.4 and from 4.9 to 9.5 respectively. The pH stability of emulsion made with the modified milk proteins was monitored by following the increase of particle size during storage. The results suggested that succinylation and EDA modification could enhanced the emulsion stability at pH 4-7 by increasing the electrostatic repulsion between droplets. Regarding the enzymatic modification of milk proteins, the laccase and transglutaminase (Tgase) catalyzed cross-linking were applied on WPI and CN respectively. In order to improve the reactivity of WPI towards the laccase, a vanillic acid modification was carried out to incorporate additional methoxyphenol groups into the protein surface. The crosslinking of vanillic acid modified WPI (Van-WPI) by laccase was studied using SDS-PAGE. The extent of cross-linking of Van-WPI was found to be significantly higher compared to the unmodified WPI and the combination of WPI and free phenolic compound as a mediator. The effect of laccase catalyzed cross-linking on storage stability was investigated by visual observation and confocal microscopy. The post-emulsification cross-linking was proven to enhance the stability of the emulsions prepared with Van-WPI during the storage. The reduced droplet coalescence could be most likely attributed to an extended interfacial protein layer formed via the interaction between the adsorbed proteins and non-adsorbed proteins in the water phase. In contrast with the limited extent of crosslinking of WPI by laccase, CN was extensively cross-linked by Tgase. The physical stability of emulsions was studied by measuring the increase of particle size during storage, and the oxidative stability was evaluated by following the formation of fatty acid hydroperoxides and volatile compounds in different stages of the lipid oxidation. The pre-emulsification cross-linking showed no obvious influence on the physical stability of CN emulsion but significantly improved its stability against lipid oxidation. The improvement of oxidative stability of emulsions could be contributed to a thicker and denser interfacial protein layer and thus increases the amount of anti-oxidative groups located at the interface and provides a stronger barrier against competitive adsorption by oil oxidation products.
|Award date||11 Sep 2015|
|Place of Publication||Espoo|
|Publication status||Published - 2015|
|MoE publication type||G5 Doctoral dissertation (article)|
- milk proteins
- protein modifications
- physical stability
- oxidative stability
Ma, H. (2015). Role of chemical and enzymatic modifications of milk proteins on emulsion stability/properties: Approaches for more stable protein emulsions: Dissertation. VTT Technical Research Centre of Finland.