Edible Oleogels for Reduction of Saturated Fat

Lead Research Organisation: Heriot-Watt University
Department Name: Sch of Energy, Geosci, Infrast & Society

Abstract

Removing saturated fats from foods is highly desirable because of the health benefits that would be realised, but this is not a trivial exercise because solids fats contribute greatly to the texture of many common foods. They are important in forming the structure in many formulated foods such as baked goods, biscuits, butter, margarine and spreads and confectionery. However, they raise cholesterol levels in the blood and are a risk factor for coronary heart disease. Replacing them with mono-or polyunsaturated oils is not feasible as these are liquid at ambient temperatures and so food where solid fat is replaced by oil would lack the desired solid texture. Even foods that are branded as high in polyunsaturated fats, such as margarines and spreads must contain a relatively high proportion of saturated fat to give the correct texture. The role of the saturated fats in spreads is to provide a network of fat crystals that trap and immobilize the liquid oils in a semi-solid matrix. If food manufacturers are to develop foods with the saturated fat removed they will need to find alternative methods to form the semi-solid structure. One solution that food scientists are developing is the use of edible oleogels. In these, molecules known as gelators are added to liquid oils to mimic the structuring effect of solid fat crystals. The gelators associate with each other to form very long, thin fibres or tubules which crosslink to form a network. This "lattice" of gelator fibres acts in similar way to the network of solid fat crystals and traps the liquid oils in pores in the structure. One group of molecules that can be used to make oleogels are the plant sterols. These also have been found to reduce blood cholesterol levels in their own right, and are added to some functional food spreads for this reason.
In principle, oleogels can have a solid-like texture the same as conventional spreads and margarines. In practice it has proven difficult to develop this technology and to date they have not been used for commercial foods. One reason is that there are few available sterols that form organogels. Since little is known about what makes the ideal gelator it is not possible to make new gelators for a particular food application. In addition the structure and texture of an oleogel is very sensitive to the presence of small amounts of water, and so margarines (which contain small water droplets) have proven difficult to make. To make matters worse, when it does prove possible to make an oleogel, it is often difficult to do so consistently. Again, little is known about how oleogels form, and this makes the control of their texture difficult.
In this project we will address the problems that are holding back the use of edible oleogels in foods. We use a combination of experimental and computer modelling methods to explore the mechanisms of gelator aggregation, tubule formation and gelation. Our aim is to understand how these molecules are able to form tubules, and subsequently how these tubules are able to form a semi-solid texture in liquid oils. Computer modelling allows us to look at the structure of tubules in molecular detail, and to understand the features of a phytosterol molecule that allows it to form an oleogel. Knowing the key structural features of optimum gelator molecules allows new gelators to be synthesised and tested in foods, leading to a wider range of sterol gelators and more widespread application to oily foods. Even with more efficient gelators the formulation of edible oleogels will be difficult. We will also look at the mechanisms of and control of gel formation using a range of experimental techniques, with the ultimate aim that we will use this knowledge to control oleogel structure, and eventually to demonstrate oleogel technology in food products. Successful formulation of edible oleogels will allow healthier oil-based foods that are reduced in saturated fats but maintain a desirable semi-solid or solid texture.

Technical Summary

Oleogels are a form of organogel where the continuous phase is unsaturated triglyceride oil trapped in a network formed by self-associating oleo-gelator molecules. Oleogels are of interest to food companies who manufacture foods such as polyunsaturated margarines and spreads. These contain saturated fat crystals added to give a semi-solid texture. Saturated fats raise blood cholesterol which is a risk factor in cardiovascular disease. Removing saturated fats from these products by using oleo-gelation would lead to a healthier product. Mixtures of sterols and sterol esters as oleogelators are of interest because they have been shown to have blood cholesterol lowering properties in their own right. There are a number of technical difficulties associated with developing oleogel food products from phytosterols. Several sterols are suitable for oleogelation, but only one sterol ester, gamma-oryzanol is available. The mechanism for self-association is not well understood, and it is not possible to predict suitable olegelators for a particular application. Oleogels made from phytosterols are sensitive to oscillatory shear and considerable super-cooling can occur in the gelation process if the shear conditions are not optimised. Finally, the self-association is sensitive to water, and this limits application of oleogelation in water-in-oil emulsion spreads and margarines. We will use a combination of experimental (rheological, AFM, light scattering) and simulation (MD, MC, Lattice-Boltzmann simulation) methodologies to (1) identify other sterol ester structures to broaden the range available; (2) understand and control the effects of shear to optimise oleogel formation and structure; and (3) understand water sensitivity and how to ensure self-association in the presence of water. This will provide information allowing us to identify the optimal routes to processing of oleogel based foods.

Planned Impact

A number of benefits, both societal and economic, have been identified that could occur as a consequence of this project. Consumption of saturated fats has been linked to an increased risk of coronary heart disease, since saturated fats increase the levels of blood cholesterol. However, solid fats are also important in the formation of structure and texture in oily foods. Replacing saturated fats with structured, polyunsaturated oil based oleogels has the potential to lead to a revolutionary change in saturated fat levels in the diet, reduction in blood cholesterol and an associated increase in health benefits. In addition to these highly significant societal benefits, the potential economic benefits are also significant. Products that contain saturated fats include spreads, chocolate, cream and ice cream. However, far greater impact on health could be realised by incorporation of oleogel technology into staple foods such as bread. As part of the follow up of this project we would look to engage with food manufacturers who would test the oleogel technology in a range of prototype food products.
Further societal benefits can be accrued through public engagement. The area of food and health is of great interest to the general public. The opportunity exists for us to connect with the public through the Edinburgh Beltane Beacon for Public Engagement in Science an organisation (www.edinburghbeltane.net). The annual Edinburgh International Science Festival offers huge scope for promoting the results of this project to a wider general audience. Both HWU and UoE are members of the Edinburgh Beltane. To reach members of the public who would not normally attend science fairs, but nonetheless have an interest in the health implications of what they eat, we would explore using other events such as the Royal Highland Agricultural show as a showcase for food and health related research.
Dissemination of research on organogels that may be interest to other academics and industrial sectors will be achieved through publication (with the prior permission of DRINC) in high impact peer-reviewed journals and presentation at international conferences. The researchers in this proposal have a track record of publication across a wide range of journal subject areas (food chemistry, physical chemistry, colloid chemistry and soft-matter physics) with this breadth of coverage ensuring outreach to a wide range of secondary beneficiaries.

The academics have the ideal fit in terms of their complementary scientific expertise, and excellent track record of engaging with industry, thus ensuring the success of the project. Euston and Clegg have a joint translational research project with industry funded by the EPSRC (EP/J501682/1 and EP/J501712/1) on novel food proteins. Euston is PI or CoI on government and industry funded projects for the food industry (TS/L002426/1, TS/L004542/1, KTP009473 & KTP009478). Clegg has been a Royal Society Industry Research Fellow with Syngenta and has contributed as PI/CoI to a number of industry related projects. He is Director of the Edinburgh Complex Fluid Partnership (ECFP, www.edinburghcomplexfluids.com): the industrial collaboration vehicle of the Edinburgh University Soft Condensed Matter Group. Existing or recent collaborations of ECFP are with AkzoNobel, Mentholatum, Johnson Matthey, Macphie of Glenbervie and Rowett Institute for Nutrition and Health. He led the Edinburgh University half of a recent DRINC collaboration with the Institute of Food Research (PI Wilde, BB/I006133/1) on the mouth feel of emulsions. Stewart has a wide portfolio of grants related to the links between food and health funded by the EU (FP7 KBBE/ 613513, KBBE/ 613793), KTP and the Scottish Government.Westacott has been or is PI or CoI on EPSRC (GR/S12005/1, EP/D003679/1, EP/G029601/1) grants based completely or partly in molecular simulation of interfaces and on KTP projects funded by the TSB (KTP000058 and KTP009119).

Publications

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Dalkas G (2018) Molecular Interactions behind the Self-Assembly and Microstructure of Mixed Sterol Organogels. in Langmuir : the ACS journal of surfaces and colloids

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Matheson AB (2017) Microstructure of ß-Sitosterol:?-Oryzanol Edible Organogels. in Langmuir : the ACS journal of surfaces and colloids

 
Description We have studied the self-association of the sterol ß-sitosterol, and the sterol ester ?-oryzanol to form tubules that are able to form solid gels in a liquid triglyceride oil. These have the potential to replace crystalline saturated fats that are used to solidify foods such as margarines and spreads. We combined both rheology and atomic force microscopy to probe the gelation mechanism to an unprecedented level. Surprisingly, we found that the oleogel fibres associated in ribbons, which did not twist. This appears to contradict previous suggestions in this area. Spectroscopic results implicate the ferulic acid group of the oryzanol in the sticking together of fibres into ribbons. Our published work has covered the classic oleogel combinations of sitosterol + ?-oryzanol, cholesterol + ?-oryzanol, cholestenol + ?-oryzanol, stigmasterol + ?-oryzanol. More recently, our gelation and rheology studies have moved on to different combinations of molecules: sitosterol + CHEMS and sitosterol with synthetic ester derivatives of sitosterol with modified versions of ferulic acid attached, all of which were found not to gel. Autodock simulations (described below) are greatly supporting these studies. This enables to test our understanding of the self-assembly behaviour in an unprecedented way.
To understand the self-association process, and to facilitate in-silico design of new sterol-ester gelators we have adapted the Autodock molecular docking program to investigate the formation of dimers as the first step in the self-association process. Autodock allows us to rapidly screen sterol-sterol ester combinations (and to identify those that form stable dimers with the potential to form oleogels.
Alternatives to oryzanol, of which the main component is cycloartenyl ferulate, cannot be identified as easily because the number of commercially available sterol esters is limited. We believe that this is due to the fact the methyl group at C30 on the cycloartenol core prevents sterane cores from stacking parallel. Limits of available materials have prevented us from testing this hypothesis. For this reason, we have synthesised and obtained a range of aromatic sterol-esters to test this hypothesis further and explore if they can behave in the same way as oryzanol. The novel aromatic esters (sitosteryl esters) did not form gels with sitosterol in sunflower oil, failing a simple vial inversion test, and the lack of gel formation strongly implies that the molecular interactions which underpin sterol-oryzanol gels are not taking place in this system. We also see no FTIR signature to indicate intermolecular hydrogen bonding between the sterol and the sterol ester, unlike in sitosterol-oryzanol blends where a new shoulder develops at 3441 cm-1 wavenumbers. Molecular docking simulations agree with this, showing that there is not a hydrogen bond forming due to the propensity for the sitosteryl cores to sit parallel on top of each other. This is qualitatively different from sterols with oryzanol. This experimental confirmation would strongly suggest that the manner these cores stack in is of key importance. To further investigate the importance of these core groups, we decided to test the gelation capabilities of blends of oryzanol with lanosterol and saponified oryzanol (the main component of which should be cycloartenol), which have the C30 methyl groups. We observe that these molecules do form transparent gels with oryzanol, but the formation mechanism seems much slower.
We have also developed a molecular dynamics model for the structure of a self-assembled tubule of and have used this to study the stability of a sitosterol+oryzanol and a sitosterol+cholesteryl hemisuccinate (CHEMS). CHEMS is a sterol ester where the ferulic acid of oryzanol is replaced with a carboxylic acid. The hydrogen bond that stabilises the sitosterol-oryzanol dimer is absent when CHEMS is substituted for oryzanol in the Autodock calculations, and is also absent in in-silico tubules. The simulations also reveal the importance of the hydrophobic interaction between the sterane cores of the sterol and sterol ester, and pi-pi (stacking) interactions between the aromatic rings of the ferulate groups attached to the oryzanol molecule. No pi-pi-stacking can occur between the CHEMS molecules in the tubule, since the ferulate group is missing from CHEMS. Experiments with CHEMS plus sitosterol reveal that the system does not gel, thus highlighting the importance of the H-bond, and possibly ferulate pi-pi interactions to tubule formation. In this respect the simulation results confirm experiments, and have allowed us to explore in greater detail the molecular origins of any interactions.
Using the tubule model, we have carried out very large scale MD simulations of two sitosterol-oryzanol tubules interacting through surface-to-surface contact to understand better the origin of the interactions that control fibril association into larger ribbon-like structures. We observed significant interactions at the surface between the two tubules that could explain the further association of tubules into ribbons. The results suggest that the ferulic acid of the ?-oryzanol may cause fibrils to stick together through hydrophobic contacts of the aromatic group of the ferulic acid moieties and through hydrogen bonds between the hydroxyl groups of the ferulic acid moieties.
To investigate the water sensitivity of the system we have carried out experiments on combining water and oryzanol within a lecithin based system, and made gels where glycerol, andalternative polar solvent, is substituted for water. Addition of lecithin in place of sitostereol permits gel formation while circumventing the historic problem of combining phytosterols with water. Specifically, rheology studies showed that by mixing these materials at an equimolar ratio, highly viscous suspensions are created. Furthermore, by adding water to these samples, a solid-like gel may be formed which offers mechanical properties close to those desired for a margarine type spread, whilst still solubilizing the oryzanol. The relationship between the oryzanol and the phospholipid molecules have been explored using both spectroscopy, Autodock and molecular dynamics simulations. Molecular dynamics simulations indicate that the mechanism for this is the formation of mixed micelles, whereby the oryzanol intercalates between lecithin molecules. Oryzanol appears to sit between the lecithin molecules in such a manner that the packing ratio of lecithin molecules is not significantly altered, and thus the micelles remain spherical. Subsequent experiments focused on controlling the composition and understanding the structure of a composite gelator system incorporating glycerol. We have demonstrated that we can develop this into an emulsion system, incorporating plant sterols, with robust gel properties. We have explored how changing the total volume of glycerol droplets, and changing the water content of these droplets alters the strength of sterol gels. We find that gels exhibit significant elastic moduli even with 30% w/w glycerol dispersed throughout the matrix. At higher glycerol loadings, we found that complex multiple emulsion morphologies can form. Molecular dynamics simulations of oryzanol+sitosterol tubules in organic solvent, water or glycerol show clearly that the cooperative hydrogen bonding network that stabilizes the tubules in organic solvents is largely absent in water, and the tubules are unstable. In glycerol, the destabilizing effect is diminished, but tubules are still not as stable as in organic solvent.
Exploitation Route The project is funded by the BBSRC Diet and Health Research Industry Club (DRINC) partly funded by the food industry. We have established contact with one of the funders, Unilever and continue to discuss the progress of this project with them. We have also had preliminary discussions with Sainsburys about potential applications for the oleogelation technology.
Computational studies will be applied to investigate the alternative candidate derivatives of oryzanol and sitosterol, and these will be synthesized for comparative studies. This will enable us to probe experimentally the roles of different chemical groups, and to identify potential gelators with improved functionality. Additionally, we are just beginning to explore the role of changing the host solvent. By this route we should be able to systematically weaken and destroy the oleogel - while observing the changes to the molecules using spectroscopic studies. This should provide some insight into how the oleogel "works". We are also beginning to investigate alternative gelator systems which can incorporate ?-oryzanol. There is an outside chance that these might prove useful for gels which need to be robust to the presence of water.
Sectors Agriculture, Food and Drink,Chemicals,Healthcare,Manufacturing, including Industrial Biotechology,Pharmaceuticals and Medical Biotechnology

URL http://www.foodproteins.org
 
Description Collaboration with Derick Rousseau 
Organisation Ryerson University
Country Canada 
Sector Academic/University 
PI Contribution Coarse-grained molecular simulation of monoglyceride self-assembly at triglyceride-water and air-water interfaces.
Collaborator Contribution Experimental studies on the molecular templating effect of monoglycerides on triglyceride crystallization at oil-water interfaces
Impact A poster and oral presentation was given at Food Colloids 2016 held at Wageningen university the Netherlands. A manuscript has been submitted and is under review.
Start Year 2016
 
Description Collaboration with Paul Clegg 
Organisation University of Edinburgh
Country United Kingdom 
Sector Academic/University 
PI Contribution This project was collaborative with UoE who provided experimental expertise to complement computer modelling at HWU.
Collaborator Contribution Provided expertise in soft matter physics, atomic force microscopy, spectroscopy.
Impact We have authored 5 papers (6th in preparation). These are listed under this award.
Start Year 2016
 
Description Organic synthesis of alternative gelator molecules 
Organisation Heriot-Watt University
Department Department of Mathematics
Country United Kingdom 
Sector Academic/University 
PI Contribution Shared information on olegelation. Invited collaborator to project meetings. Eventual joint publications.
Collaborator Contribution The partner will synthesize a range of alternative olegelator molecules that are derivatives of gamma oryzanol that either have the ferullic acid side chain substituted for another group, or have a hydroxyl group substituted. The hydroxyl group is believed to be important in dimerization of the sterols prior to their subsequent self-association into tubules, and also stabilizes the tubules through cooperative H-bonding. The ferrulic acid group is believed to allow inter-tubule interaction (it sticks out from the surface of the tubule) via pi-pi stacking interactions, thus leading to gel formation. By creating and testing oleogelators where one or both of these groups has been substituted will allow us to understand the role they play in the self-association and gelation process.
Impact Too early in project/collaboration.
Start Year 2017
 
Description Attendance at Diet and Health Research industry Club Industry engagement meetings 
Form Of Engagement Activity A formal working group, expert panel or dialogue
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Industry/Business
Results and Impact Mandatory DRINC dissemination event to inform industry sponsors of the DRINC program of the progress and outcomes of DRINC funded research project. Three have been attended (1 per year) so far.
Year(s) Of Engagement Activity 2015,2016,2017
 
Description Conference paper at Physics in Food manufacturing event, Chipping Campden, January 2019. 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact Workshop organised by the Institute of Physics food group. Allowed networking with physicists/physical chemists with similar intersts in food science.
Year(s) Of Engagement Activity 2019
 
Description Conference paper on Food Oleogels at EFFost conference November 2018 in Nantes France 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Discussion were held with industrial (and academic) scientists who were interested in the finding of the work.The computational approaches used were of particular interests to academics and this has fostered new interactions with potential academic collaborators.
Year(s) Of Engagement Activity 2018
 
Description Food Colloids Conference, Wageningen The Netherlands 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Results presented at a major food colloids conference (sponsored by the Royal Society of Chemistry) held in Wageningen, The Netherlands in April 2016.
A poster was given on the computer simulation of olegelation by sterols and sterol esters, and on the functional properties of novel fungal proteins.
Year(s) Of Engagement Activity 2016
 
Description Seminar at University of Leeds 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Other audiences
Results and Impact Seminar on edible oleogels at the University of Leeds to academic staff and students
Year(s) Of Engagement Activity 2017