Sampson Anankanbil

Adding functionality to milk-based protein: Preparation, and physico-chemical characterization of β-lactoglobulin-phenolic conjugates

Multi-functional phenolic emulsifiers were prepared by covalently coupling β-Lactoglobulin (βLg) to caffeic acid (CA) using crosslinker chemistry at different pH conditions (pH 2.5, 6.0, and 8.5). The resulting bioconjugates were characterized by MALDI-TOF MS, differential scanning calorimetry (DSC), fluorescence-quenching, infrared and circular dichroism spectroscopies. Furthermore, the emulsifying and antioxidant properties of βLg-CA conjugates were evaluated and compared to native β-Lactoglobulin and the non-covalent β-lactoglobulin/caffeic complex (βLg/CA). Results showed: 1) An optimal molar ratio (8:1) of caffeic acid to βLg was obtained at pH 6; 2) DPPH activity of βLg-CA increases as the number of CA units coupled increases; 3) βLg-CA conjugates displayed comparable or superior water solubility than native βLg and βLg/CA. Moreover, DSC results showed that coupling of CA with βLg significantly increased the thermal stability of βLg. In summary, βLg-CA conjugates can act as effective antioxidant emulsifiers and stabilizers and may find application in food and cosmetic industries.

A novel array of interface-confined molecules: Assembling natural segments for delivery of multi-functionalities

HYPOTHESIS Anionic surfactants can form stable monolayers around oil droplets via interactions with macromolecules thereby physically stabilizing high fish oil enriched emulsions (50-70% fish oil) while phenolic acids have antioxidant properties to prevent lipid oxidation. COSMO-RS (Conductor-like Screening Model for Real Solvents) is a powerful tool for the rational design of molecules with multi-functionalities. Therefore, it should be possible to assemble segments of natural molecules into a single multifunctional molecule using COSMO-RS to confer both physical and oxidative stability to fish oil enriched systems. EXPERIMENTS COSMO-RS was used to predict the thermodynamic properties of series phenoleoyl malic acid esters of monoglycerides in comparison with commercial emulsifiers. A novel series of amphiphilic lipids, equipped with multi-functional groups from natural building blocks (fatty-acyl, glycerol, malic & phenolic acids), were then synthesized in a facile approach and characterized by various spectroscopy techniques. Oil-in-water emulsions stabilized by the amphiphilic lipids were formulated and characterized by dynamic light scattering measurements and fluorescence imaging. FINDINGS An elaborate integration of multi-functions into a single molecule was achieved, displaying superior or comparable emulsion stability and antioxidant property, compared to a commercial emulsifier, phenolic acids and their combinations. This is the first report to holistically integrate the rational design, synthesis and functional characterization of natural-based multifunctional molecules for high capacity fish oil delivery systems.

Enzymatic alkylsuccinylation of tyrosol: Synthesis, characterization and property evaluation as a dual-functional antioxidant

This work reports a novel approach to generate a new group of tyrosol-based amphiphilic lipid alkylsuccinylated tyrosol by lipase-catalyzed succinylation of tyrosol with alkylsuccinic anhydrides of varying alkyl chain lengths, in high yields (80-95%). The structures of the compounds were confirmed by MS, FTIR &1H NMR; and their properties were characterized by Temperature-Ramp FTIR, DSC & CMC measurements. The synthesized compounds integrate water-soluble phenylethanoid and hydrophobic alkyl into one molecule thus are endowed with dual functions: retaining the antioxidant property of tyrosol and entailing tyrosol with new surface-active property. The DPPH activity of tyrosol (13.77%) was significantly enhanced by 2-dodecen-1-ylsuccinylated tyrosol (16.01%). Compared to tyrosol-based emulsions (76.63%), the lipid oxidation is reduced to 21.57% and 42.32% in 2-octen-1-ylsuccinylated/2-dodecen-1-ylsuccinylated tyrosol emulsions, respectively. This work brings new members to the library of functional lipid excipients and open a novel and effective synthetic pathway for derivation of phenyl alcohols.

Combination of sodium caseinate and succinylated alginate improved stability of high fat fish oil-in-water emulsions

Sodium caseinate (CAS) and commercial sodium alginate (CA), long chain modified alginate (LCMA) or short chain modified alginate (SCMA) were used in combination for emulsifying and stabilizing high fat (50-70%) fish oil-in-water emulsions. Physical (creaming, droplet size, viscosity and protein determination) and oxidative (primary and secondary oxidation products) stabilities of the emulsions were studied during 12 days of storage. Creaming stability was higher for emulsions produced with alginates and CAS compared to emulsions prepared with only CAS. Combined use of CAS + LCMA performed better in terms of physical stability compared to emulsions produced with only CAS. However, the oxidative stability of this emulsion was inferior probably due to the presence of an unsaturated carbon chain in LCMA structure. CAS + SCMA emulsions not only showed better physical stability such as smaller droplet size, lower creaming and higher viscosity, but also had an improved oxidative stability than emulsions produced with only CAS.

Valorizing dairy waste: thermophilic biosynthesis of a novel ascorbic acid derivative

l-Ascorbic acid (l-AA) is an essential nutrient that is extremely unstable and cannot be synthesized by the human body. Therefore, attempts have been performed to develop biologically active l-AA derivatives with improved stability. This work presents a facile, scalable, and efficient enzymatic transgalactosylation of lactose to l-AA using β-glucosidase (TN0602) from Thermotoga naphthophila RKU-10. β-Glucosidase TN0602 displays high transgalactosylation activity at pH 5.0, 75 °C, and l-AA/lactose ratio of 2:1 to form a novel l-AA derivative [2-O-β-d-galactopyranosyl-l-ascorbic acid (l-AA-Gal)] with a maximal productivity of 138.88 mmol L-1 in 12 h, which is higher than most reports of enzymatic synthesis of l-AA-α-glucoside. Synthetic l-AA-Gal retains most l-AA antioxidant capability and presents dramatically higher stability than l-AA in an oxidative environment (Cu2+). In conclusion, this work reports a new way to valorize dairy waste lactose into a novel molecule l-AA-Gal, which could be a promising l-AA derivative to be used in a wide range of applications.

Synthesis of Sugar Fatty Acid Esters and Their Industrial Utilizations

Sugar fatty acid esters (SFAEs) are widely used as nontoxic biosurfactants. This chapter reviews the latest progress in the development of novel enzymatic and chemical synthetic approaches to obtain classic and novel SFAEs. Furthermore, it presents the updated knowledge about physical properties of SFAEs and new advances in their applications in the pharmaceutical, cosmetic, and food industry.

Caffeoyl maleic fatty alcohol monoesters: Synthesis, characterization and antioxidant assessment

HYPOTHESIS Caffeoyl malate anhydride, as a good nucleophilic acceptor, can react with lipophilic fatty alcohols to yield interface-confined amphiphiles. The resulting novel molecules are hypothesized to deliver combined functionalities of parent natural building blocks, as emulsifier, stabilizer, ion chelator and free radical scavenger. EXPERIMENTS Ring-opening reactions of caffeoyl malate anhydride with fatty alcohols of different chain lengths generated a new group of antioxidant amphiphiles. Structural verification was by MS (mass spectrometry), 1H/13C NMR (nuclear magnetic resonance) and FT-IR (Fourier transform infra-red) spectroscopy. Physicochemical characterization was done by use of DSC (differential scanning calorimetry), FT-IR, determinations of critical micelle concentrations (CMC) and calculations of HLB. Antioxidant activity was assessed by DPPH (2, 2-diphenyl-1-picrylhydrazyl) and hydroxyl radical scavenging activities. Dynamic light scattering (DLS) studies demonstrated surface-activity of G8-G18. Inhibition of iron- and thermally-accelerated lipid oxidation was monitored by thiobarbituric acid reactive substances (TBARS) assay. FINDINGS Derivatization of caffeoyl malate anhydride with fatty alcohols maintained free radical scavenging activity, and improved hydroxyl radical scavenging activity of caffeic acid. Lipid oxidation at 22 °C was significantly inhibited (up to 3.5 times) in emulsions stabilized by G8-G18 with or without chitosan compared to emulsions stabilized by commercial emulsifiers and stabilizers. Thermal oxidation (at 80 °C) was 10 times less in emulsions facilitated by G8-G18 in combination with chitosan compared to emulsions stabilized by commercial emulsifiers and stabilizers. This study has developed a simple and straightforward approach for developing value-added compounds from underexplored fatty alcohols.

Mapping the location of DATEM in multi-phase systems: Synthesis and characterization of spin-label probe analogues

Electron paramagnetic resonance (EPR) spectroscopy has emerged as a fast, reliable, non-invasive and sensitive method to determine the distribution, localization and reactivity of labelled ingredients in micro-heterogeneous systems. However, the commercially available probe molecules are very limited. In the present work, five new nitroxide [(4-hydroxy-2, 2, 6, 6-tetramethylpiperidin-1-oxyl (TEMPOL)] derivatives (1b-5b) of diacetyl tartaric acid esters of monoglycerides (DATEM) (1a-5a), a widely used food emulsifier, were synthesized under Steglich conditions and characterized by MS (mass-spectrometry), FT-IR (Fourier-transform infrared spectroscopy), EPR, NMR (nuclear magnetic resonance spectroscopy), fluorescence spectroscopy, and DSC (differential scanning calorimetry) analysis. Phase partitioning studies proved that the new spin labels are adequately capable of describing the localizations and partitioning of the corresponding DATEM in multi-phase systems. Findings disclosed in this work will provide new knowledge on ingredient reactivity and localization in multi-phase systems; which is vital to aid the design of more efficient delivery systems for bioactive compounds.

A new group of synthetic phenolic-containing amphiphilic molecules for multipurpose applications: Physico-chemical characterization and cell-toxicity study

Nine synthetic amphiphilic phenolic lipids, varied in phenolic moiety (caffeoyl/dimethylcaffeoyl) and fatty acid chain lengths (8–18) were characterized by differential scanning calorimetry (DSC), temperature-ramp Fourier transform infra-red spectroscopy (FT-IR) and atomic force microscopy (AFM). FT-IR and DSC results revealed that the physical state and lateral packing of synthetic molecules were largely governed by fatty acyls. The critical micelle concentrations (CMC) of synthetic lipids was in the range of 0.1 mM to 2.5 mM, affording generation of stable oil-in-water emulsions; as evidenced by the creaming index (<5%) of emulsions stabilized by compounds C12‒C16, and C12a‒C16a after 7 days’ storage. AFM analysis revealed that compound C14 formed stable double-layers films of 5.2 nm and 6.7 nm. Application studies showed that formulations stabilized by synthesized compounds containing 30% fish oil had superior physical and oxidative stability compared to formulations containing commercial emulsifiers or their mixtures with phenolic acids. Moreover, the synthetic compounds were non-toxic against in vitro transformed keratinocytes from histologically normal skin and Caco-2 cell lines. This study demonstrates the relevance of using a natural hydroxycarboxylic acid as a flexible linker between natural antioxidants, glycerol and fatty acids to generate multifunctional amphiphiles with potential applications in food, pharmaceutical and cosmetic industry.

New phenophospholipids equipped with multi-functionalities: Regiospecific synthesis and characterization

HYPOTHESIS In multi-phase systems, many complex reactions take place at the interface where a molecule equipped with manifold functionalities is demanded. By taking advantage of the surface-active property of phosphatidylcholine (PC) scaffold and antioxidant properties of phenolic acids, new multifunctional molecules are generated, which are expected to confer physical and oxidative stability to sensitive bioactive ingredients in delivery systems. EXPERIMENTS This work reports a successful synthesis of two new arrays of phenophospholipids sn-1-acyl(C12-C18)-sn-2-caffeoyl and sn-1-caffeoyl-sn-2-acyl phosphatidylcholines via mild scalable regiospecific pathways; as structurally verified by MS, 1H/13C NMR analyses, and characterized by critical micelle concentrations (CMC), FTIR, and DSC analysis. Synthesized phenophospholipids are subjected to stabilizing o/w emulsion, and antioxidation tests as demonstrated by TBARS (Thiobarbituric Acid Reactive Substances) and DPPH (2,2-diphenyl-1-picrylhydrazyl) assays. FINDINGS This study has demonstrated that; (1) phenophospholipids with a broad spectrum of CMC are created, affording superior emulsion stability than soybean PC; (2) all phenophospholipids present improved oxidation inhibition and sn-2-caffeoyl phenophospholipids display superior performance to sn-1-caffeoyl phenophospholipids, soybean PC or admixture of caffeic acid and soybean PC; (3) incorporation of caffeoyl in PC scaffold does not sacrifice radical scavenging ability of caffeic acid, whilst the ion chelating capacity of sn-1-myristoyl(C14)-sn-2-caffeoyl PC enhance by 4.5 times compared to soy PC. Fluorescence Microscopy imaging verified the location of phenophospholipids in the interface as desired. Among synthetic phenophospholipids, sn-1-myristoyl(C14)-sn-2-caffeoyl PC commits the cut-off effect in most desired functionalities, which might be of great potential for multi-purpose applications.