Background of the invention Biopolymers are extensively used in food products as emulsifiers, gelling agent, structuring agents, stabilisers, thickeners and the like. Especially biopolymers such as starches, protein and triglycerides are well known and are used for example in sauces, margarine, dressings, soups and many other food compositions.

Many of the polymers are naturally occurring and show good functionality in relation to the above mentioned features.

However there is still a need for alternative compositions with improved functionality and controlled characteristics. This desire is exemplified in WO-A-96/03440, which discloses a method for gelling or increase of viscosity of aqueous media containing gellable polymeric materials having substituents with phenolic hydroxyl groups. This document specifically discloses the gelation of arabinoxylans or pectins by adding an effective amount of a laccase to an aqueous medium containing these substituents. Furthermore it is disclosed that proteins having one or more tyrosine residues in the amino acid sequence can be gelled by use of a laccase. The resulting compounds are indicated as thickening and/or stabilising agents.

The compositions disclosed in WO-A-96/03440 rely on the gelation of single compositions and hence lead to gels with a limited variety in composition and functionality.

It is therefore an object of the current invention to provide polymers which can be used in a variety of food products and which enlarge the group of currently available polymers for use in foods. The invention especially relates to polymers suitable for stabilising oil and water containing emulsions and foams.

Summary of the invention It has surprisingly been found that specific polymers of a glyceride and a molecule (M) selected from the group of protein, polysaccharides and/or glycerides meet the above objective. Especially those polymers consisting of a polysaccharide and a fatty acid glyceride unit were found to be good emulsifiers and/or stabilisers.

Therefore the invention relates to a polymer composition comprising a glyceride and a molecule (M) selected from the group comprising proteins (P), glycerides (G) and polysaccharides (S), wherein the molecule (M) and the glyceride are covalently linked via phenolic residues forming polymer building blocks of P-G, G-G, G-S or combinations thereof.

The invention further relates to food products containing these polymers and to a method for preparation of these polymers.

Detailed description of the invention

The polymers according to the invention are composed of glycerides and molecules (M) which together form building blocks. Building blocks are polymer units which comprise a glyceride and a molecule (M) selected from the group comprising glycerides, proteins and polysaccharides. The glyceride and molecule (M) are covalently linked by 2 connected phenolic residues. This covalent bond is an essential part of the claimed polymers. The individual building blocks may be linked together via further covalent bonds, preferably between 2 phenolic residues.

The building blocks of the polymers according to the invention comprise glycerides and at least one of polysaccharides (S), glycerides (G) and proteins (P).

Glycerides in the context of the invention are glycerol based molecules wherein the glycerol backbone is covalently linked to at least one residue such as a fatty acid, or another hydrophobic acid, aldehyde or ketone e. g. retinoic acid, phenolics, sterols, tocopherols. Well known glycerides are mono, di and tri acyl fatty acid glycerides. Mono-and diglycerides are already known for their emulsifying capacity.

Triglycerides are well known texturing agents in products such as margarine, butter, creams.

The glycerides may contain any fatty acid, both naturally occurring and synthetic ones.

Preferred fatty acids have a chain length of from 10 to 24 carbon atoms, more preferred from 16 to 20 carbon atoms.

Suitable fatty acids include oleic acid, stearic acid, palmitic acid linoleic and linolenic acid.

Most preferred, the glyceride in the building block is a diglyceride.

Molecule (M) optionally is a protein. Proteins are chains of covalently linked amino acids whereby it is possible to use any desired protein. Preferred proteins contain at least one lysine amino acid residue. Food grade proteins are preferred and hence protein is preferably selected from the group of dairy proteins such as casein and whey protein, vegetable proteins, especially soy protein and egg protein or gelatin.

The third potential molecule of a building block is polysaccharide. Any polysaccharide can be used in the polymers according to the invention but preferred are those polysaccharides which in their naturally occurring form contain a phenolic residue such as ferulic acid, vannilic acid, coumaric or cinnamic acid esterified to (M). Sugar beet pectin and arabinoxylanes isolated from cereals are examples of polymers that contain ferulic acid residues in their naturally occurring form. These compositions are described in more detail by Lex Oosterveld, Carbohydrate research 300,179-181, 1997 and thesis Lex Oosterveld, Landbouwuniversiteit Wageningen Netherlands, 16.12. 1997, ISBN 90-5485785-4.

The glyceride and molecule (M) in the polymer comprise a link via two phenolic residues. This link is a covalent bond. It is preferred that the phenolic residue that is participating in the covalent bond comprises a radical stabilising group in ortho position in the phenolic ring. The substituent is preferably such that the resulting compound is food grade. An example of a very suitable electron-withdrawing group is the methoxygroup, positioned ortho with respect to the hydroxyl group.

Examples of covalent bonds between two phenolic residues are described by Oosterveld (thesis Lex Oosterveld 1997) and in US 5,786, 470. Preferably the phenolic residue is derived from the group of vanillic acid, ferulic acid, coniferol, caffeic acid, chlorogenic acid.

Figure 1 is one example of a specific covalent bond between the two phenolic residues. Other forms of covalent bonding are also encompassed within the invention.

The polymer according to the invention comprises a glyceride and at least another molecule selected from protein, glyceride, polysaccharide or a combination thereof. Embodiments wherein n building blocks are linked to obtain a large linear polymer are also encompassed within the invention. Further encompassed in the invention are polymers wherein building blocks are linked via at least 2 phenolic residues or by other (non) covalent linkages. The polymers according to the invention may be both linear and branched polymers.

In the polymers the covalent linkage is between P-G, G-G, G-S, or combinations thereof.

These alternative embodiments are described below.

In a first embodiment, the polymer comprises covalently linked proteins and glycerides, preferably diglycerides. These polymers when branched form fat-protein networks which may be applied as structuring agents.

In another embodiment the polymer comprises covalently linked polysaccharides and glycerides, preferably diglycerides. An example of such a composition is shown in figure 1. These compositions were found to be suitable emulsifiers which may be used to stabilise oil and water containing compositions. In

such compositions the polymers are found at the oil/water interface whereby the fatty acid chains are in the oil phase and the polysaccharide part is in the aqueous phase. Preferred glycerides for this purpose are diglycerides containing fatty acids derived from sunflower oil, soybean oil, rapeseed oil, maizgerm oil, olive oil, line oil, peanuts, cottonseed oil, and safflower oil, butter fat.

Preferred polysaccharides for this purpose are selected from the group comprising pectins from sugar beet, arabinoxylanes, starches. These polysaccharides may naturally comprise a phenolic group or may have been functionalised with such group.

According to a further embodiment the polymer comprises covalently linked glycerides. The glycerides may be mono or diglycerides with any suitable fatty acid. These polymers preferably contain a multiplicity (n) of glyceride building blocks, with a preference for n between 2 and 1000.

Such polymers may be used to structure a fat phase for example in a margarine type food composition.

Preferred polymers are those wherein a glyceride is covalently linked to a polysaccharide or protein. Most preferred compositions are those wherein a glyceride is covalently linked to a polysaccharide because these polymers were found to have emulsion stabilising properties.

The polymers according to the invention comprise at least 2 phenolic ring systems. Surprisingly these were found to enhance the stability of unsaturated fatty acid residues that may be part of the polymer and of other oxidation sensitive compositions in the same product. Without wishing to be bound by any theory, applicants believe this is due to the mesomeric

ring system which can trap radicals which would otherwise cause oxidation of the double carbon-carbon bond.

A As indicated above, the polymers according to the invention may suitably be applied in food products, whereby depending on their composition the properties of the food product may be influenced. Their use is especially recommended in oil and water containing compositions.

Therefore in a further aspect the invention relates to food products containing said polymers.

The following beneficial characteristics of these compositions have been identified.

(G-G) n which are referred to as acyl-glycerol-phenolic acid hybrids, have superior emulsifying properties especially in oil and water containing compositions of relatively low fat content (i. e. between 10 and 60 wt% fat). Furthermore these compositions may be used as texturising agents, imparting viscosity and/or structure to a composition.

Furthermore these compositions, especially (G-G) n (n equal to or higher than 1) containing polymers are suitable for encapsulation of ingredients such as flavour compounds or functional ingredients. To make such (G-G) n compositions, it is preferred to start from monoglycerides functionalised with two phenolic acid groups.

Polymers containing (G-S) n wherein n is preferably 1, were found to stabilise oil and water containing compositions.

Furthermore these compositions when included in a two phase system such as water and oil, can fix particles or micro- organisms in one of the phases.

Therefore in a further aspect, the invention relates to an oil and water containing emulsion, preferably an oil in water emulsion, comprising a polymer according to invention.

In a preferred embodiment, the invention relates to an oil and water emulsion comprising from 20 to 80 wt% fat and from 0.1 to 10 wt% of said polymer. Even more preferred, the polymer comprises covalently linked glyceride and polysaccharide.

The polymers according to the invention are prepared from glycerides and molecule (M) under suitable circumstances to form a covalent bond. In a further aspect the invention relates to a method for the preparation of said polymer, wherein a composition comprising the elements of the building blocks is treated with an oxidative enzyme. This treatment may suitably be carried out in presence or absence of a solvent. For most cases, the use of an aqueous medium is preferred. However for example in case of the preparation of a glyceride-glyceride hybride polymer, no solvent is needed as long as the oxidation enzyme is functional. The latter can for example be obtained by using well known immobilisation methods for the enzyme.

Before oxidation takes place the molecules (building block elements) that form the starting composition in this reaction are preferably functionalised with one or more phenolic residues, preferably containing an ortho methyxygroup with respect to the hydroxyl group. It will be appreciated that functionalisation is not needed for molecules that naturally contain phenolic residues.

Said functionalisation may be obtained through natural occurrence of said phenolic groups in the element of a building

block or by synthetic functionalisation of said phenolic groups to the respective molecule. It will be appreciated that 4 naturally occurring polymers with phenolic groups are preferred.

In the oxidation reaction the phenolic groups are covalently linked. An example of a possible covalent link is shown in figure 1.

The oxidation may be carried out in any way known in the art.

Both enzymatic oxidation and chemical oxidation routes may be used. Enzymatic oxidation is preferred. Suitable enzymes that can catalyse the formation of the covalent bond are peroxidase, laccase, polyphenol oxydases. Peroxidase is the preferred enzyme.

In case an enzymatic oxidising system is applied, the enzyme is preferably added in the form of a solution or a dispersion in an aqueous buffer system.

Some enzymes, such as peroxidases, require the presence of a co-oxidant such as hydrogen peroxide for their activity. The co-oxidant is preferably added separately from the enzyme that requires it's presence.

The amount of enzyme added is expressed in terms of activity units. Preferably enzyme is present in excess.

If the oxidation is carried out enzymatically, the temperature during oxidation is preferably from 20 to 60 °C. Most preferred the temperature is around the temperature at which the enzyme shows maximum activity.

In case a chemical oxidant is applied, the oxidant is preferably added in the form of a diluted aqueous solution. ri The introduction of a phenolic residue is described below for each of the potential elements of building blocks.

Polysaccharides Well known esterification conditions may be used for introduction of a phenolic group on the polysaccharide chain.

Protein Some proteins naturally contain amino acids with phenolic residues. The preparation of a protein containing phenolic residues is straightforward. Any possible covalent link e. g. via esterification is suitable.

Glycerides Di/mono-glycerides containing at least one phenolic residue are preferably prepared by interesterification of a triglyceride oil with the ester form of the phenolic residue.

Examples of the latter are ferulic acid ester with para position esterified to an alcohol, vanillic acid ester with para position esterified to an alcohol or caffeic acid ester with para position esterified to an alcohol.

Said interesterification reaction can be carried out with chemical or enzymatic catalysts. The use of lipase as catalyst is preferred.

In a preferred embodiment the invention relates to a method wherein a polymer comprising covalently linked diacylglyceride and polysaccharide is prepared, comprising the steps of a) preparing a feruloyl diglyceride by transesterification of

triacylglycerol with a phenolic compound containing a methoxygroup in ortho position with respect to the hydroxyl group ; b.) providing a polysaccharide composition containing a phenolic hydroxygroup containing a methoxygroup in ortho position ; c) subjecting the composition according to (a) and (b) to treatment with an oxidative enzyme.

It will be appreciated that the exact reaction conditions in terms of temperature, ratio between the elements of the building blocks and type/amount of catalyst/enzyme, determine the final product composition in terms of polymer length and composition.

The invention is illustrated by the following examples.

Examples Example 1 Synthesis of feruloylated diglyceride (DG-F) DG-F was synthesised following the method disclosed by Compton (JAOCS, vol 77, no 5,2000, pages 513-519) preparing feruloyl diolein starting from triolein.

The synthesis was performed without the use of solvent.

Ethylferulate (3.58 g) was dissloved in 40 g triolein at 60°C.

Novozyme 435 (3.55 g) (a lipase) was added and the reaction was allowed to proceed for 7 days at 60°C under continuous mixing.

Purification

5.3 g of the reaction mixture was brought onto a Silicagel 60 (Merck) column (3.5 cm diam. , 25 cm height). The mixture was subsequently separated by elution with solvent mixes with increasing polarity in the following order: petroleum ether : diethyl ether = 4: 1 (v/v), petroleum ether : diethyl ether = 7: 3 (v/v), petroleum ether : diethyl ether = 3 : 2 (v/v) and finally by elution with diethyl ether. Fractions of approximately 200ml were collected, solvent was removed by evaporation and analysis was done by means of thin layer chromatography on Kieselgel 60 F254 plates (Merck) using a mixture of toluene: diethyl ether = 4: 1 (v/v).

Emulsion tests for controls 11.85 g sunflower oil, 3 ml water and 0.15g of either Admul Woltm, (comparative example) total transesterification mix (comparative example) or purified DG-F (comparative example) were mixed and emulsified by vigorous stirring for approximately one minute.

No emulsifying properties were observed for purified DG-F or the total transesterification mix. The phases started to separate within two hours standing at room temperature.

Crosslinked polymer according to the invention In situ preparation of emulsifier: crosslinking of DG-F and pectin-F with laccase to yield DG-F-F-pectin.

A number of test tubes were filled as given in the table.

The conditions of each experiment are outlined in table 1.

Tube 1-7 and Tube 10-12 are control experiments, which are not according to the invention. Tubes 8,9 are according to the invention.

All tubes were prepared at room temperature.

First the tubes were stirred vigorously on a Vortex mixer for 30 seconds. u Next the enzyme was added (or not) and mixing was done for another 30 seconds.

Tubes were left at RT and photographed.

Table 1 Tube Gram mg pL uL UL Laccase number Sunflow DG-F water 5 % 1% er oil pectin pectin 1 1. 69-429--yes 2 1. 69 - 286 143 - yes 3 1. 69-286-143 yes 4 1. 69-429--no 5 1. 69-286 143 no 6 1. 69-286-143 no 7 1. 69 21.4 429 - - yes 8 1. 69 21. 4 286 143-yes 9 1. 69 21.4 286-143 yes 10 1. 69 21.4 429 - - no 11 1. 69 21.4 286 143 no 12 1. 69 21.4 286 - - no

Pectin = Kelco pectin in water (w/v) Enzyme = Novo Lacase PPL, SP710, PX 5326, 10 uL of a 1: 10 dilution in water was added.

Stable emulsions without phase separation of the oil and water phase were only obtained when DG-F and pectin was present and a crosslinking enzyme was used. This confirms that the polymer comprising diglyceride linked to pectin via a ferulic acid- ferulic acid covalent bond is a good emulsifier for oil/water systems.