School of Chemistry - Theses
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ItemThe effects of ultrasound on the molecular, structural and nutritional aspects of dairy proteins( 2021)Process improvement and product improvement in the dairy processing industry is an ongoing dynamic process. Ultrasound processing technology for food processing is in its developmental stages, and dairy sector is a key area for potential applications. The use of low frequency and high frequency ultrasound to modify functional properties of dairy streams is being widely studied by seeking to modify the protein and lipid components of milk. The effects of ultrasound on size of lipid globules and lipid oxidation volatiles have been studied. The effects on functional properties of milk proteins such as viscosity, gelation, heat stability etc. have been documented. However, the present literature lacks information on the effects that ultrasound processing may have on the fundamental properties of dairy proteins. Therefore, it is of importance to document the effects of ultrasound processing on the molecular, structural and nutritional aspects of dairy proteins. As such, the effects of ultrasound processing on dairy proteins have been studied in the context of the amino acid composition, amyloid modifications, and bioactive peptides released. This work aims to bridge the gap in existing literature attempting to illuminate upon these fundamental aspects. Chapter 1 in this thesis introduces the premise of this research by discussing the fundamental properties ultrasound technology and presents dairy proteins as the central theme of this research. A systematic review of literature in Chapter 2 attempts to build familiarity with the effects of processing on milk proteins; and discusses the potential applications of ultrasound in the dairy industry – noting the studies done on whey are caseins. This chapter also discusses the research areas which could be addressed to answer questions related to the objectives of current PhD work. To encourage the application of ultrasound for dairy processing, it would be of significance to understand the extent of operating parameters that ensure nutritional integrity and food safety. The amino acid integrity of skim milk proteins after sonication along with the potential of ultrasound to modify secondary structural conformations of milk proteins has been demonstrated in Chapter 4. Chapter 5 expands the objectives of Chapter 4 into whole milk systems, taking into account milk lipids and the potential for oxidation of lipids on sonication. In Chapter 5 an effort has been made to better understand the underlying cause of lipid oxidation observed due to ultrasound processing of milk lipids. The observations from this study suggest that the enhanced mass transfer of oxygen due to ultrasonic shear forces makes oxygen non-limiting in lipid phase and increases auto-oxidation reactions. An assessment of amino acid integrity of full cream milk after ultrasound treatment confirmed oxidative stability of milk proteins. These findings complement the data in Chapter 4. Along with amino acid integrity, Chapter 4 also demonstrates that the secondary structural conformations of milk proteins can be modified by ultrasound. The work in Chapter 6 builds up greater understanding on such changes by studying the effects of low frequency ultrasound on isolated beta-lactoglobulin (b-lg). b-lg is a b-sheet rich protein with amyloidogenic potential. The effects of high shear and high temperature microenvironments produced by ultrasonic waves were studied on b-lg aggregation. Ultrasound induced formation of amyloid crystals in dilute b-lg solutions at neutral pH and ambient bulk temperatures. The theoretical reasoning for the aggregation phenomenon substantially adds to the understanding of protein energy landscape. Chapter 7 focusses on nutritional properties of milk proteins. It examines the effect of ultrasound pre-treatment on the release of BCM-7 bioactive peptide from milk by simulated gastro-intestinal digestion (SGID), and the effect on overall protein digestibility. It demonstrates that ultrasound processing could be used to downregulate the release of undesirable BCM-7 bioactive peptide in milk, by affecting the rate of enzymatic hydrolysis of certain intermediate peptide fragments. The findings from this body of work add to the existing knowledge for improved use of ultrasound to ensure nutritional integrity of dairy proteins and lipids, minimising oxidative damage. It also builds on current understanding of ultrasound induced protein restructuring and its potential for modification of peptide release from dairy proteins.
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ItemUltrasound driven synthesis of bio-functional nanostructures( 2019)Polyphenolic-, amino acids- and doxorubicin-based nanostructures are of great interest due to their multifarious applications in biomedical field as antibiotics, antioxidants, antimicrobial and anti-cancer agents. Most of the research on the polyphenolic and amino acids based nanostructures adhere to the formation of coordination complexes with metals, self-assembly techniques or chemical functionalization and crosslinking reactions. To improve the efficacy of therapeutic agents, a variety of nanoparticles have been developed for the controlled and targeted delivery of doxorubicin. These include biopolymers-based nano/microcapsules, carbon-based nanoparticles, polymer-drug conjugate, dendrimers, liposomes, micelles, inorganic nanoparticles and nucleic acids nanostructures. The development of simple, one pot and effective synthesis routes to fabricate bio-nanomaterials is in high demand. In particular, the use of polyphenols, doxorubicin and single amino acids as building blocks to fabricate nanostructured materials is still unexplored. In this PhD work, I have used ultrasound-based technologies to synthesize phenolic, amino acid and doxorubicin based micro and nanoparticles for different biomedical applications. Chapter 1 provides an overview on the structural and bio-functional properties of nanoparticles and methods to synthesize nanoparticles for biomedical use, including the ultrasonic techniques have been discussed. Furthermore, fundamentals of ultrasound are also provided. In the literature review (Chapter 2), several studies dealing with the polyphenol, doxorubicin and amino acid molecules have been summarised. In the last section of this chapter, numerous investigations on synthesis of diverse types of nanostructures using ultrasound are reviewed. In Chapter 3 materials and methods, equipment and all other experimental details used are described. Chapter 4 provides a fundamental understanding on the ultrasonic coupling of simple phenolic molecules, where acoustic bubble acts as a catalytic binding site for the generation of bioactive oligomers without the need for utilizing any enzymes, catalysts (organic or inorganic) and other toxic reagents. It has been observed that the extent of oligomerization and nanoparticles formation depends on the ultrasonic frequency, concentration and physiochemical properties of the phenolic building blocks. Chapter 5 demonstrates that cavitation bubbles are simple micro-reactors with reactive surfaces to perform one-pot multiple reactions on complex polyphenolic molecules to convert tannic acid to ellagic acid, namely (i) hydrolysis of an ester linkage, (ii) C–C coupling reactions, (iii) condensation reactions and (iv) crystallization of the product into regularly shaped particles. The size and shape of the crystals can be controlled by ultrasonic frequency, power and time. The synthesized particles exhibit fluorescence properties, anticancer and antioxidant activity and could be further used for drug loading and delivery. In Chapters 6 and 7, the role of the acoustic field in the formation of supramolecular nanoaggregates using tryptophan and phenylalanine as building blocks was investigated. It has been demonstrated that the acoustic bubbles driven at high frequency standing wave, in addition to provide a reactive surface for the dimerization of biomolecules, can also provide an energy source to fuel and refuel the dissipative out of equilibrium assembly of these molecules below the critical aggregation concentration. Furthermore, the unique optical and bio-functional properties of nanoparticles for bioimaging and probing the intracellular trafficking of a drug have also been studied. In Chapter 8 the sound-driven self-assembly of the anticancer drug doxorubicin was investigated to generate nanoparticles solely made of drug molecules. The newly developed nanoparticles were tested on different types of cancer cells and the drug was found to be active in drug resistant cell lines. In addition, the mechanism of action of drug nanoparticles was investigated. Chapter 9 provides a summary of the entire PhD work.