Investigation of Solid and Liquid State Milk Protein-Carbohydrate Interactions in Tea Blends

The development of stable and functional milk-tea powders presents a complex formulation challenge, primarily due to the component interactions among milk proteins, carbohydrates, lipids, and polyphenolic compounds. These interactions depend heavily on the formulation and are highly sensitive to processing steps such as pasteurisation, concentration, drying, and storage. Each stage can disrupt the structural stability of the systems and reduce its functional performance. As a result, conventional milk-tea products often experience poor phenolic retention and powder instability, which shorten the shelf life and lower antioxidant capacity. To address these limitations, this research systematically investigated how varying the lactose-to-maltodextrin (L:M) and casein-to-whey (C:W) protein ratios, along with tea infusion concentration, influenced the physicochemical and structural behaviours of fat-filled (FM-T) and skim (SM-T) milk-tea systems across liquid, concentrated, and powder states. The primary goal was to optimise compositional balance and reveal the underlying molecular mechanisms responsible for stabilisation, with the ultimate aim of developing shelf-stable milk-tea powders with enhanced functional quality. The study first examined the effect of L:M variation on the physicochemical and structural properties of FM-T and SM-T during pasteurisation and concentration (Chapter 3). Optimal L:M ratios (90:10 and 75:25 for FM-T; 90:10 and 80:20 for SM-T) were selected based on their capacity to enhance total phenolic content (TPC), reduce particle size, improve zeta potential, and control lactose crystallisation. In FM-T, these formulations achieved up to 160 % improvement in TPC, attributed to the formation of protective fat-carbohydrate-polyphenol matrices and stable electrostatic environments. Although SM-T demonstrated improved dispersion stability and structural properties at similar L:M ratios, phenolic retention remained comparatively lower due to the absence of fat, which otherwise provides a hydrophobic microenvironment essential for polyphenol entrapment. These findings indicated that while maltodextrin contributes to improved matrix characteristics, the synergistic presence of fat is critical for optimal phenolic protection. To support the matrix and protein-polyphenol complexation, the formulations were further optimised by adjusting the C:W ratio (Chapter 4). The inclusion of whey protein (up to 30 %) significantly enhanced the surface charge and TPC retention, particularly in SM-T, where whey balanced for the lack of fat by forming disulfide-stabilised and hydrogen-bonded polyphenol complexes. In FM-T, the synergy among fat, maltodextrin, and whey protein promoted a denser protein–polyphenol network, further stabilised by β-sheet structures as revealed by FTIR and SDS-PAGE. Native PAGE also confirmed increased protein aggregation in FM-T, supporting the hypothesis that optimising L:M and C:W ratios can be used to modulate protein interactions and enhance phenolic retention under heat stress. Building upon these optimised L:M and C:W combinations, tea concentration was increased from 1 % to 5 % to evaluate the modifications of the systems under elevated polyphenol load (Chapter 5). Both FM-T and SM-T exhibited tea concentration-dependent increases in zeta potential after pasteurisation, attributed to hydrogen bonding and proton release from polyphenols. However, following concentration, a decline in zeta potential was observed, particularly in SM-T, due to polyphenol oversaturation leading to excessive protein-polyphenol aggregation. FM-T benefited from fat-mediated hydrophobic interactions that preserved secondary structure and limited disulfide bonding, while SM-T showed increased aggregation, disulfide cross-linking, and larger particle sizes, especially in high-casein systems. Volatile analysis further indicated that FM-T retained a more complex aroma profile dominated by lipid oxidation and Maillard products, while SM-T exhibited a simpler aldehyde- and alcohol-based profile. These results emphasised that while tea enrichment can enhance antioxidant capacity, it also introduces destabilisation risks, especially in fat-free systems lacking adequate protein-polyphenol protecting mechanisms. Chapter 6 focused on spray-dried powders and their performance during storage under varying relative humidity (RH) and temperatures. Distinct stabilisation mechanisms emerged depending on composition. In FM-T, fat-driven lipid migration during spray drying contributed to surface oxidation but improved dispersibility and hydrophobic barrier formation. Increasing maltodextrin (75:25 L:M) enhanced carbohydrate–lipid interactions and steric hindrance, stabilising the amorphous phase and increasing thermal resistance. However, fat still hindered strong protein–polyphenol cross-linking, which limited network rigidity. In SM-T, polyphenol–protein interactions dominated. A 90:10 L:M and 80:20 C:W formulation promoted disulfide-linked whey protein networks with high thermal stability but poor dispersibility. In contrast, SM-T with 80:20 L:M and 70:30 C:W formed highly rigid, protein–carbohydrate–polyphenol matrices with excellent oxidation resistance. The final phase (Chapter 7) provided a molecular-level understanding of the mechanisms underlying the powder behaviour and stability across formulations with different L:M and C:W ratios. Using a comprehensive set of analytical techniques, key structural and functional differences between FM-T and SM-T were revealed. X-ray Photoelectron Spectroscopy (XPS) confirmed that spray drying led to distinct surface compositions where FM-T powders exhibited fat-enriched surfaces due to lipid migration, while SM-T showed protein-enriched surfaces. Fourier Transform Infrared Spectroscopy (FTIR) identified differences in protein secondary structures and cross-linking patterns, with FM-T displaying more extensive lipid–protein–carbohydrate associations and SM-T showing stronger polyphenol–protein interactions. PAGE (native and SDS-PAGE) demonstrated that SM-T had higher disulfide-linked aggregation of β-Lg, BSA, and IgG, indicating enhanced protein–polyphenol binding in the absence of fat. Differential Scanning Calorimetry (DSC) showed that SM-T had higher glass transition temperatures (Tg), attributed to its higher protein content and lower plasticizer load, while X-ray Diffraction (XRD) confirmed that lactose remained largely amorphous. Finally, Small-Angle X-ray Scattering (SAXS) showed that FM-T possessed looser micellar arrangements due to fat-induced disruptions, whereas SM-T maintained compact casein networks stabilized by calcium–phosphate interactions. Collectively, these findings illustrated that compositional differences govern the internal microstructure, interfacial behaviour, and thermal strength of milk-tea powders. In conclusion, this study demonstrates that the structural and functional stability of milk-tea powders can be finely controlled through rational optimisation of L:M and C:W ratios. The synergistic effects of fat, maltodextrin, and whey proteins allowed for the design of modified matrices that withstand thermal and storage stresses while preserving antioxidant functionality. FM-T formulations offered superior dispersion, flavour complexity, and polyphenol protection under thermal stress, while SM-T achieved higher glass transition temperatures and microstructural stability through protein–polyphenol cross-linking. However, SM-T remained more vulnerable to phenolic overload, emphasizing the importance of balancing tea concentration and matrix design. This study provides a mechanistic framework for designing next-generation milk-tea powders with extended shelf life, improved rehydration behaviour, and optimised health-promoting attributes.<p></p>