EP3774757A1

EP3774757A1 - Chroman-6-ols with an extended lipophilic side chain in position 2, their manufacture and use

Info

Publication number: EP3774757A1
Application number: EP19714655.8A
Authority: EP
Inventors: Laure CLASADONTE; André DUESTERLOH; Weerasinghe INDRASENA; Thomas Netscher; René Tobias STEMMLER
Original assignee: DSM IP Assets BV
Current assignee: DSM IP Assets BV
Priority date: 2018-03-29
Filing date: 2019-03-29
Publication date: 2021-02-17
Also published as: CN113166090A; WO2019185904A1

Abstract

The present invention is directed to a process for the manufacture of a compound of formula (III), comprising the step of reacting a compound of formula (I) with a compound of formula (II) in the presence of an acid catalyst and in a mixture of two solvents, wherein n is 1 or 2, and R¹ and R³ are independently from each other H or C_1-5-alkyl, and R² is either H or C_1-5-alkyl or C_1-5-alkyloxy, preferably with the proviso that at least one of R¹, R² and R³ is H, and wherein the first of the two solvents is selected from ethylene carbonate, propylene carbonate, 1,2-butylene carbonate, gamma-butyrolactone and water, and the second of the two solvents is selected from hexane, cyclohexane, heptane, ortho-xylene, meta-xylene, para-xylene, mesitylene, pseudocumene, methyl tert-butyl ether, and toluene. The present invention is also directed to the compound of formula (III-3), as well as to the use of compounds of formula (III), especially to the use of compound of formula (III-3) and the use of compound of formula (III-4), as antioxidants, especially in feed, such as e.g. in feed for aquatic animals, in feed for terrestrial animals (especially pet food, feed for pigs and feed for poultry) and in feed for insects, and in feed ingredients, such as in fish meal, in poultry meal, in insect meal, and in PUFA-containing oils such as marine oil, microbial oil, fungal oil, algal oil and PUFA-containing plant oil The present invention is further directed towards feed and feed ingredients containing at least one compound of formula (III), especially containing a compound of formula (III-3) and/or a compound of formula (III-4).

Description

Chroman-6-ols with an extended lipophilic side chain in position 2. their manufacture and use

Summary

The present invention is directed to the compound of formula (MI-3), as well as to the use of compounds of formula (III) as defined below, especially to the use of compound of formula (111-3) and the use of compound of formula (111-4), as antioxidants, especially in feed, such as e.g. in feed for aquatic animals, in feed for terrestrial animals (especially pet food, feed for pigs and feed for poultry) and in feed for insects, and in feed ingredients, such as in fish meal, in poultry meal, in insect meal, and in PUFA-containing oils such as marine oil, microbial oil, fungal oil, algal oil and PUFA-containing plant oil.

The present invention is further directed towards feed and feed ingredients containing at least one compound of formula (III) as defined below, especially containing a compound of formula (MI-3) and/or a compound of formula (MI-4).

The present invention is also directed to a process for the manufacture of a compound of formula (III), comprising the step of reacting a compound of formula (I) with a compound of formula (II) in the presence of an acid catalyst and in a mixture of two solvents,

wherein n is 1 or 2, and

R¹ and R³ are independently from each other H or Ci-₅-alkyl, and R² is either H or Ci-₅-alkyl or Ci-₅-alkyloxy, preferably with the proviso that at least one of R¹, R² and R³ is H,

the first of the two solvents is selected from ethylene carbonate, propylene carbonate, 1 ,2-butylene carbonate, gamma-butyrolactone and water,

and the second of the two solvents is selected from hexane, cyclohexane, heptane, ortho-xylene, meta-xylene, para-xylene, mesitylene, pseudocumene, methyl tert- butyl ether, and toluene.

Until now there was no sustainable industrial process for the manufacture of compounds of formula (III) with a good yield based on the compound of formula (I), whereby the use of halogenated solvents is avoided. Thus, there was a need to provide such a process.

Detailed description of the invention Thus, this need is fulfilled by the present invention, which is directed to a process for the manufacture of a compound of formula (III),

comprising the step of reacting a compound of formula (I) with a compound of formula (II) in the presence of an acid catalyst and in a mixture of two solvents,

( )

wherein n is 1 or 2, and

R¹ and R³ are independently from each other H or Ci-5-alkyl, and R² is either H or Ci-5-alkyl or Ci-5-alkyloxy, preferably with the proviso that at least one of R¹, R² and R³ is H,

The compound of formula (111-3), 2-(4,8-dimethylnonyl)-2-methylchroman-6-ol, is a novel compound and therefore also an object of the present invention.

The two asterisks ^* mark each a chiral/stereogenic center. The term“compound of formula (111-3)” encompasses all possible isomers having any configuration at said centers.

Unmodified fish meal can spontaneously combust from heat generated by oxidation of the polyunsaturated fatty acids in the fish meal. In the past, factory ships have sunk because of such fires. Strict rules regarding the safe transport of fish meal have been put in place by authorities and the International Maritime Organization (IMO). According to IMO, fishmeal must be stabilized with antioxidants to prevent spontaneous combustion during overseas transport and storage.

The shipping regulations of the United Nations for the Transport of Dangerous Goods (UN-TDG) currently only allow ethoxyquin and BHT as antioxidants to stabilize fish meal for marine transport. But authorization of ethoxyquin has now been suspended in the European Union due to safety and health concerns.

BHT must be added in higher quantities to achieve the same efficacy as ethoxyquin. Furthermore, BHT is currently under safety evaluation by ECHA and its re-registration as feed additive is pending in Europe.

Therefore, there is a need to replace ethoxyquin and BHT as an antioxidant, especially in feed comprising proteins and/or unsaturated fatty acid (derivative)s and in feed ingredients comprising proteins and/or unsaturated fatty acid

(derivative)s. Such replacements are the compounds of formula (III), especially the compound of formula (111-3) as well as the compound of formula (MI-4).

Thus, the compounds of formula (III), especially the compound of formula (MI-3) as well as the compound of formula (MI-4), are useful as antioxidants, especially in feed, such as e.g. in feed for aquatic animals, in feed for terrestrial animals (especially pet food, feed for pigs and feed for poultry) and in feed for insects, and in feed ingredients, such as in fish meal, in poultry meal, in insect meal, and in PUFA-containing oils such as marine oil, microbial oil, fungal oil, algal oil and PUFA-containing plant oil. Thus, the present invention is further directed to this use.

The asterisk ^* marks a chiral/stereogenic center. The term“compound of formula (111-4)” encompasses all possible isomers having any configuration at said center.

Furthermore, the present invention is directed towards feed and feed ingredients containing at least one compound of formula (III), especially containing a compound of formula (111-3) and/or a compound of formula (MI-4).

Feed encompasses feed for aquatic animals, feed for terrestrial animals (especially pet food, feed for pigs and feed for poultry) and feed for insects. Feed ingredients encompass especially fish meal, poultry meal, insect meal, and PUFA-containing oils such as marine oil, microbial oil, fungal oil, algal oil and PUFA-containing plant oil.

“PUFA(s)” means polyunsaturated fatty acid(s) such as docosahexaenoic acid (“DHA”) and/or eicosapentaenoic acid (“EPA”) and/or docosapentaenoic acid (“DPA”) and/or oleic acid and/or stearidonic acid and/or linoleic acid and/or alpha- linolenic acid (“ALA”) and/or gamma-linolenic acid and/or arachidonic acid (“ARA”) and/or the esters of all of them, whereby the term “esters” encompasses monoglycerides, diglycerides and triglycerides as well as Ci-₆-alkyl esters such as especially the methyl esters and the ethyl esters, whereby the triglycerides are often dominant. DHA, EPA, ALA and stearidonic acid are omega-3 fatty acids, whereas linoleic acid, gamma-linolenic acid and ARA are omega-6 fatty acids.

The term“DPA” encompasses two isomers, the omega-3 fatty acid clupanodonic acid (7Z,10Z,13Z,16Z,19Z-docosapentaenoic acid) and the omega-6 fatty acid osbond acid (4Z,7Z, 10 Z, 13Z, 16Z-docosapentaenoic acid).

In accordance with the invention, the polyunsaturated fatty acid (PUFA) is preferably DHA and/or EPA and/or DPA and/or any ester thereof, more preferably the polyunsaturated fatty acid (PUFA) is preferably DHA and/or EPA and/or any ester thereof.

Examples of PUFA-containing oils are

- marine oil, such as preferably fish oil,

- microbial biomass containing polyunsaturated fatty acids and/or their esters

(“microbial oil”), preferably containing high amounts of docosahexaenoic acid (“DHA”) and/or eicosapentaenoic acid (“EPA”) and/or

docosapentaenoic acid (“DPA”) and/or their esters, and

- oil containing high amounts of PUFAs and/or their esters, preferably

containing high amounts of docosahexaenoic acid (“DHA”) and/or

eicosapentaenoic acid (“EPA”) and/or docosapentaenoic acid (“DPA”) and/or their esters, extracted from microbial biomass, such as fungae

(“fungal oil”) or algae (“algal oil”), and

- plant oil with relatively high amounts of PUFAs and/or their esters, (“PUFA- containing plant oil”), such as e.g. canola seed oil, linseed /flaxseed oil, hempseed oil, pumpkin seed oil, evening primrose oil, borage seed oil, blackcurrent seed oil, sallow thorn/sea buckthorn oil, chia seed oil, argan oil and walnut oil. Thus, in addition, the present invention is

(1 ) directed to the use of the compounds of formula (III) as antioxidants in feed, such as especially feed for aquatic animals, feed for terrestrial animals such as poultry, pigs and pets, and feed for insects; as well as (2) directed to the use of the compounds of formula (III) as antioxidants in feed ingredients, such as especially poultry meal, fish meal, insect meal and PUFA- containing oil, and

(3) directed to feed, such as especially feed for aquatic animals, feed for terrestrial animals such as poultry, pigs and pets, and feed for insects, comprising such compounds of formula (III) and

(4) directed to feed ingredients, such as especially poultry meal, fish meal, insect meal and PUFA enriched oil, comprising such compounds of formula (III). Thus, the present invention is directed to feed for aquatic animals comprising such compounds of formula (III) with the preferences as given above.

The present invention is also directed to feed for insects and terrestrial animals, e.g. pigs, poultry and pets, comprising such compounds of formula (III) with the preferences as given above.

Aquatic animals in the context of the present invention encompass farmed

Crustacea such as shrimp and carnivorous species of farmed fish such as salmons, rainbow trout, brown trout (Salmo trutta) and gilthead seabream.

Thus, the feed for aquatic animals comprising the compounds of formula (III) are especially fed to the aquatic animals as cited above.

I. Feed ingredients

Feed ingredients are broadly classified into cereal grains, protein meals, fats and oils, minerals, feed additives, and miscellaneous raw materials, such as roots and tubers.

Further antioxidants

The compounds of formula (III) can be used in combination with one or more other antioxidants as described below. In an embodiment of the present invention the feed ingredients of the present invention additionally comprise a mixture of 2-tert-butyl-4-methoxyphenol and 3- tert-butyl-4-methoxyphenol, which is known under the name “BHA” (butylated hydroxyanisole).

In a further embodiment of the present invention the feed ingredients of the present invention additionally comprise ascorbyl palmitate. In another embodiment of the present invention the feed ingredients of the present invention additionally comprise BHA and ascorbyl palmitate.

Instead of ascorbyl palmitate other esters of ascorbic acid such as the esters of ascorbic acid with linear C_12-20 alkanols, preferably the esters of ascorbic acid with linear C_14-18 alkanols, may also be used, so that further embodiments of the present invention are directed to feed ingredients that additionally comprise esters of ascorbic acid with linear C_12-20 alkanols, preferably esters of ascorbic acid with linear C_14-18 alkanols, more preferably ascorbyl palmitate, whereby optionally BHA may also be present.

The feed ingredients may also comprise additionally alpha-tocopherol and/or gamma-tocopherol, whereby either an ester of ascorbic acid with a linear C_12-20 alkanol with the preferences as given above or BHA or both may additionally be present.

The feed ingredients themselves are described in more detail below.

1 . PUFA-containing oils

In the context of the present invention the term“PUFA-containing oil”

encompasses

- marine oil, such as especially fish oil,

- microbial biomass containing polyunsaturated fatty acids (“PUFAs”), especially docosahexaenoic acid (“DHA”) and/or eicosapentaenoic acid (“EPA”) and/or docosapentaenoic acid (“DPA”) and/or their esters (“microbial oil”);

- oil containing high amounts of PUFAs, especially containing high amounts of DHA and/or EPA and/or DPA and/or their esters extracted from microbial biomass as e.g. , fungi (“fungal oil”) or algae (“algal oil”);

- Plant oil with high amounts of PUFAs and/or their esters (“PUFA-containing plant oil”), such as e.g. canola seed oil, linseed /flaxseed oil, hempseed oil, pumpkin seed oil, evening primrose oil, borage seed oil, blackcurrent seed oil, sallow thorn/sea buckthorn oil, chia seed oil, argan oil and walnut oil.

The term“DHA” does not only encompass the acid but also derivatives thereof such as monoglycerides, diglycerides and triglycerides as well as Ci-₆-alkyl esters such as the methyl and ethyl esters. The same applies for“EPA” and“DPA” and all the other PUFAs.

Fish oil and algal oil are common feed ingredients. Instead of fish oil and algal oil also the other PUFA-containing oils named above may be used as feed ingredients, i.e.:

- microbial biomass containing PUFAs (“microbial oil”)

- oil containing high amounts of PUFAs extracted from microbial biomass, such as especially fungal oil, and

- plant oil with high amounts of PUFAs.

The above-mentioned feed ingredients may not only be used as alternative of fish oil and algal oil, but also in addition.

Examples of PUFA-containing oils that are used as feed ingredients are given below in more detail. Marine oil

Examples of suitable marine oils include, but are not limited to, Atlantic fish oil, Pacific fish oil, or Mediterranean fish oil, or any mixture or combination thereof. In more specific examples, a suitable fish oil can be, but is not limited to, pollack oil, bonito oil, pilchard oil, tilapia oil, tuna oil, sea bass oil, halibut oil, spearfish oil, barracuda oil, cod oil, menhaden oil, sardine oil, anchovy oil, capelin oil, herring oil, mackerel oil, salmonid oil, tuna oil, and shark oil, including any mixture or combination thereof.

Other marine oils suitable for use herein include, but are not limited to, squid oil, cuttle fish oil, octopus oil, krill oil, seal oil, whale oil, and the like, including any mixture or combination thereof.

For stabilizing marine oil an amount of at least one compound of formula (III) ranging from 10 to 500 ppm, preferably ranging from 30 to 300 ppm, more preferably ranging from 100 to 250 ppm, based on the total amount of the marine oil, is usually sufficient. The same applies for the other PUFA-containing oils such as microbial oil, algal oil, fungal oil and PUFA-containing plant oil.

A commercially available example of marine oil is the fish oil“MEG-3” (Bleached 30S TG Fish oil) from DSM Nutritional Products, LLC (US) whose specification and composition is shown in Tables I and II below:

Table I

The peroxide value is defined as the amount of peroxide oxygen per 1 kilogram of oil. Traditionally this is expressed in units of milliequivalents or meq/kg. Winterization is part of the processing of fish oil, and it is performed to remove solid fat in the oil. The“cold test” is performed to check if any solid fat is present and precipitated in the oil when cooled to 0°C within a specific period of time. In this fish oil (Product Code: FG30TG), any such precipitation is checked for 3 hours at 0°C.

Table

“TG” = triglyceride;

“A%” =“area %” = area percentage by GC based on 24 peak analysis (meaning the 24 highest peaks have been analyzed)

Oil containing high amounts of PUFAs, especially containing high amounts of DHA and/or EPA and/or DPA and/or their esters, extracted from microbial biomass as e.g., fungi (“fungal oil”) or algae (“algal oil”)

Algal oil

“Algal oil” is an oil containing high amounts of DHA and/or EPA and/or DPA and/or their esters extracted from algae as microbial source/biomass.

An example of algal oil is the commercially available“Algal oil containing EPA+DPA” from DSM Nutritional Products, LLC (US) whose composition is shown in the Table III below:

Table III

Fatty Acid Profile

A further example of a crude oil containing high amounts of DHA and/or EPA extracted from microbial sources as e.g., algae, is the oil extracted from Algae Schizochytrium Biomass, whose specification is given in the following Table IV.

Table IV

Microbial biomass containing polyunsaturated fatty acids (“PUFAs”), especially docosahexaenoic acid and/or eicosapentaenoic acid and/or docosapentaenoic acid (“DPA”) and/or their esters The biomass preferably comprises cells which produce PUFAs heterotrophically. According to the invention, the cells are preferably selected from algae, fungi, particularly yeasts, bacteria, or protists. The cells are more preferably microbial algae or fungi.

Suitable cells of oil-producing yeasts are, in particular, strains of Yarrowia, Candida, Rhodotorula, Rhodosporidium, Cryptococcus, Trichosporon and

Lipomyces.

Oil produced by a microorganism or obtained from a microbial cell is referred to as “microbial oil”. Oil produced by algae and/or fungi is referred to as an algal and/or a fungal oil, respectively.

As used herein, a "microorganism" refers to organisms such as algae, bacteria, fungi, protist, yeast, and combinations thereof, e.g., unicellular organisms. A microorganism includes but is not limited to, golden algae (e.g., microorganisms of the kingdom Stramenopiles); green algae; diatoms; dinoflagellates (e.g., microorganisms of the order Dinophyceae including members of the genus

Crypthecodinium such as, for example, Crypthecodinium cohnii or C. cohnii); microalgae of the order Thraustochytriales; yeast ( Ascomycetes or

Basidiomycetes ); and fungi of the genera Mucor, Mortierella, including but not limited to Mortierella alpina and Mortierella sect, schmuckeri, and Pythium, including but not limited to Pythium insidiosum.

In one embodiment, the microorganisms of the kingdom Stramenopiles may in particular be selected from the following groups of microorganisms: Hamatores, Proteromonads, Opalines, Developayella, Diplophrys, Labrinthulids,

Thraustochytrids, Biosecids, Oomycetes, Hypochytridiomycetes, Commotion, Reticulosphaera, Pelagomonas, Pelagococcus, Ollicola, Aureococcus, Parmales, Diatoms, Xanthophytes, Phaeophytes (brown algae), Eustigmatophytes,

Rophidophytes, Synurids, Axodines (including Rhizochromulinales, Pedinellales, Dictyochales), Chrysomeri dales, Sarcinochrysidales, Hydrurales, Hibberdiales, and Chromulinales. In one embodiment, the microorganisms are from the genus Mortierella, genus Crypthecodinium, genus Thraustochytrium, and mixtures thereof. In a further embodiment, the microorganisms are from Crypthecodinium Cohnii. In a further embodiment, the microorganisms are from Mortierella alpina. In a still further embodiment, the microorganisms are from Schizochytrium sp. In yet an even further embodiment, the microorganisms are selected from Crypthecodinium Cohnii, Mortierella alpina, Schizochytrium sp., and mixtures thereof.

In a still further embodiment, the microorganisms include, but are not limited to, microorganisms belonging to the genus Mortierella, genus Conidiobolus, genus Pythium, genus Phytophthora, genus Penicillium, genus Cladosporium, genus Mucor, genus Fusarium, genus Aspergillus, genus Rhodotorula, genus

Entomophthora, genus Echinosporangium, and genus Saprolegnia.

In an even further embodiment, the microorganisms are from microalgae of the order Thraustochytriales, which includes, but is not limited to, the genera

Thraustochytrium (species include arudimentale, aureum, benthicola, globosum, kinnei, motivum, multirudimentale, pachydermum, proliferum, roseum, striatum); the genera Schizochytrium (species include aggregatum, limnaceum, mangrovei, minutum, octosporum); the genera Ulkenia (species include

amoeboidea, kerguelensis, minuta, profunda, radiate, sailens, sarkariana, schizochytrops, visurgensis, yorkensis); the genera Aurantiacochytrium; the genera Oblongichytrium; the genera Sicyoidochytium; the genera Parientichytrium; the genera Botryochytrium; and combinations thereof. Species described within Ulkenia will be considered to be members of the genus Schizochytrium. In another embodiment, the microorganisms are from the order Thraustochytriales. In yet another embodiment, the microorganisms are from Thraustochytrium. In still a further embodiment, the microorganisms are from Schizochytrium sp.

In certain embodiments, the oil can comprise a marine oil. Examples of suitable marine oils are the ones as given above.

The biomass according to the invention preferably comprises cells, and preferably consists essentially of such cells, of the taxon Labyrinthulomycetes ( Labyrinthulea , net slime fungi, slime nets), in particular, those from the family of Thraustochytriaceae. The family of the Thraustochytriaceae ( Thraustochytrids ) includes the genera Althomia, Aplanochytrium, Aurantiochytrium, Botryochytrium, Elnia, Japonochytrium, Oblongichytrium, Parietichytrium, Schizochytrium, Sicyoidochytrium, Thraustochytrium, and Ulkenia. The biomass particularly preferably comprises cells from the genera Aurantiochytrium, Oblongichytrium, Schizochytrium, or Thraustochytrium, more preferably from the genus

Schizochytrium.

In accordance with the invention, the polyunsaturated fatty acid (PUFA) is preferably DHA and/or EPA and/or their esters as defined above.

The cells present in the biomass are preferably distinguished by the fact that they contain at least 20 weight-%, preferably at least 30 weight-%, in particular at least 35 weight-%, of PUFAs, in each case based on cell dry matter.

In a very preferred embodiment of the current invention, cells, in particular a Schizochytrium strain, is employed which produces a significant amount of EPA and DHA, simultaneously, wherein DHA is preferably produced in an amount of at least 20 weight-%, preferably in an amount of at least 30 weight-%, in particular in an amount of 30 to 50 weight-%, and EPA is produced in an amount of at least 5 weight-%, preferably in an amount of at least 10 weight-%, in particular in an amount of 10 to 20 weight-% (in relation to the total amount of lipid as contained in the cells, respectively). Preferred species of microorganisms of the genus Schizochytrium, which produce EPA and DHA simultaneously in significant amounts, as mentioned before, are deposited under ATCC Accession No. PTA-10208, PTA-10209, PTA-10210, or PTA- 1021 1 , PTA-10212, PTA-10213, PTA-10214, PTA-10215. DHA and EPA producing Schizochytrium strains can be obtained by consecutive mutagenesis followed by suitable selection of mutant strains which demonstrate superior EPA and DHA production and a specific EPA:DHA ratio. Any chemical or nonchemical (e.g. ultraviolet (UV) radiation) agent capable of inducing genetic change to the yeast cell can be used as the mutagen. These agents can be used alone or in combination with one another, and the chemical agents can be used neat or with a solvent.

Methods for producing the biomass, in particular, a biomass which comprises cells containing lipids, in particular PUFAs, particularly of the order Thraustochytriales, are described in detail in the prior art (see e.g. WO 91 /07498, WO 94/08467, WO 97/37032, WO 97/36996, WO 01 /54510). As a rule, the production takes place by cells being cultured in a fermenter in the presence of a carbon source and a nitrogen source, along with a number of additional substances like minerals that allow growth of the microorganisms and production of the PUFAs. In this context, biomass densities of more than 100 grams per litre and production rates of more than 0.5 gram of lipid per litre per hour may be attained. The process is preferably carried out in what is known as a fed-batch process, i.e. the carbon and nitrogen sources are fed in incrementally during the fermentation. When the desired biomass has been obtained, lipid production may be induced by various measures, for example by limiting the nitrogen source, the carbon source or the oxygen content or combinations of these.

In a preferred embodiment of the current invention, the cells are grown until they reach a biomass density of at least 80 or 100 g/l, more preferably at least 120 or 140 g/l, in particular at least 160 or 180 g/l (calculated as dry-matter content). Such processes are for example disclosed in US 7,732,170.

Preferably, the cells are fermented in a medium with low salinity, in particular, so as to avoid corrosion. This can be achieved by using chlorine-free sodium salts as the sodium source instead of sodium chloride, such as, for example, sodium sulphate, sodium carbonate, sodium hydrogen carbonate or soda ash. Preferably, chloride is used in the fermentation in amounts of less than 3 g/l, in particular, less than 500 mg/l, especially preferably less than 100 mg/l.

PUFA-containing plant oils: Plant oils with relatively high amounts of PUFAs. especially with high amounts of DHA and/or EPA such as e.g. , canola seed oil The plant cells may, in particular, be selected from cells of the families

Brassi caceae , Elaeagnaceae and Fabaceae. The cells of the family Brassicaceae may be selected from the genus Brassica, in particular, from oilseed rape, turnip rape and Indian mustard; the cells of the family Elaeagnaceae may be selected from the genus Elaeagnus, in particular, from the species Oleae europaea ; the cells of the family Fabaceae may be selected from the genus Glycine, in particular, from the species Glycine max.

Examples:

- Canola seed oil with a content of DHA of at least 9% by weight, of at least

12% by weight, of at least 15% by weight, or of at least 20% by weight, based on the total weight of the canola seed oil;

- Canola seed oil with a content of EPA of at least 9% by weight, of at least 12% by weight, of at least 15% by weight, or of at least 20% by weight, based on the total weight of the canola seed oil.

Examples of PUFA-containing plant oils containing high amounts of other PUFAs than EPA and/or DHA and/or DPA and/or their esters are linseed /flaxseed oil, hempseed oil, pumpkin seed oil, evening primrose oil, borage seed oil, blackcurrent seed oil, sallow thorn/sea buckthorn oil, chia seed oil, argan oil and walnut oil.

2. Other feed ingredients Poultry meal/Chicken meal

Poultry meal is a high-protein commodity used as a feed ingredient. It is made from grinding clean, rendered parts of poultry carcasses and can contain bones, offal, undeveloped eggs, and some feathers. Poultry meal quality and composition can change from one batch to another.

Chicken meal, like poultry meal, is made of "dry, ground, rendered clean parts of the chicken carcass" according to AAFCO and may contain the same ingredients as poultry meal. Chicken meal can vary in quality from batch to batch. Chicken meal costs less than chicken muscle meat and lacks the digestibility of chicken muscle meat.

Poultry meal contains preferably not less than 50 weight-% of crude protein, not less than 5 weight-% of crude fat, not more than 5 weight-% of crude fiber, not more than 40 weight-% of ash and not more than 15 weight-% of water, each based on the total weight of the poultry meal, whereby the total amount of all ingredients sums up to 100 weight-%. More preferably poultry meal contains from 50 to 85 weight-% of crude protein, and from 5 to 20 weight-% of crude fat, and from 1 to 5 weight-% of crude fiber, and from 5 to 40 weight-% of ash, and from 5 to 15 weight-% of water, each based on the total weight of the poultry meal, whereby the total amount of all ingredients sums up to 100 weight-%.

For stabilizing poultry meal an amount of at least one compound of formula (III) ranging from 10 to 1000 ppm, preferably ranging from 30 to 700 ppm, more preferably ranging from 100 to 500 ppm, based on the total amount of the poultry meal, is usually sufficient.

The same amounts also apply for chicken meal.

Fish meal

Fish meal contains preferably not less than 50 weight-% of crude protein, and not more than 20 weight-% of crude fat, and not more than 10 weight-% of crude fibers, and not more than 25 weight-% of ash, and not more than 15 weight-% of water, each based on the total weight of the fish meal, whereby the total amount of all ingredients sums up to 100 weight-%. More preferably fish meal contains from 50 to 90 weight-% of crude protein and from 5 to 20 weight-% of crude fat, and from 1 to 10 weight-% of crude fibers, and from 5 to 25 weight-% of ash, and from 5 to 15 weight-% of water, each based on the total weight of the fish meal, whereby the total amount of all ingredients sums up to 100 weight-%. For stabilizing fish meal an amount of at least one compound of formula (III) ranging from 10 to 2000 ppm, preferably ranging from 100 to 1500 ppm, more preferably ranging from 300 to 1000 ppm, based on the total amount of the fish meal, is usually sufficient.

Fish meal is a commercial product made from fish that is used primarily as a protein supplement in compound feed, especially for feeding farmed fish, Crustacea, pigs and poultry, and companion animals such as cats and dogs.

A portion of the fish meal is made from the bones and offal left over from processing fish used for human consumption, while the larger percentage is manufactured from wild-caught, small marine fish. It is powder or cake obtained by drying the fish or fish trimmings, often after cooking, and then grinding it. If the fish used is a fatty fish it is first pressed to extract most of the fish oil.

The uses and need of fish meal are increasing due to the rising demand for fish, because fish has the best feed conversion rate of all farmed animals, can be produced well in developing countries and has a small size, i.e. can be slaughtered for preparing a meal, so that there is no need to store the fish. Furthermore, there are no religious constraints concerning the consumption of fish, fish is a source of high quality protein and it is easy to digest.

Fish meal is made by cooking, pressing, drying, and grinding of fish or fish waste to which no other matter has been added. It is a solid product from which most of the water is removed and some or all of the oil is removed. About four or five tons of fish are needed to manufacture one ton of dry fish meal.

Of the several ways of making fish meal from raw fish, the simplest is to let the fish dry out in the sun. This method is still used in some parts of the world where processing plants are not available, but the end-product is of poor quality in comparison with ones made by modern methods.

Currently, all industrial fish meal is usually made by the following process: Cooking: A commercial cooker is a long, steam-jacketed cylinder through which the fish are moved by a screw conveyor. This is a critical stage in preparing the fishmeal, as incomplete cooking means the liquid from the fish cannot be pressed out satisfactorily and overcooking makes the material too soft for pressing. No drying occurs in the cooking stage.

Pressing: A perforated tube with increasing pressure is used for this process. This stage involves removing some of the oil and water from the material and the solid is known as press cake. The water content in pressing is reduced from 70% to about 50% and oil is reduced to 4%.

Drying: If the fish meal is under-dried, moulds or bacteria may grow. If it is over- dried, scorching may occur and this reduces the nutritional value of the meal.

The two main types of dryers are:

Direct: Very hot air at a temperature of about 500 °C is passed over the material as it is tumbled rapidly in a cylindrical drum. This is the quicker method, but heat damage is much more likely if the process is not carefully controlled.

Indirect: A cylinder containing steam-heated discs is used, which also tumbles the meal.

Grinding: This last step in processing involves the breakdown of any lumps or particles of bone.

The fish meal has to be transported long distances by ship or other vehicles to the various locations, where it is used.

Unmodified fish meal can spontaneously combust from heat generated by oxidation of the polyunsaturated fatty acids in the fish meal. Therefore, it has to be stabilized by antioxidants. Especially advantageous for this purpose are the compounds of formula (III) of the present invention.

Insect meal Insect meal has a high content of protein and is therefore, a valuable source of protein.

In general any insect may be manufactured to meal, but insects of special interest in the context of the present invention encompass black soldier flies (Hermetia species, commonly called BSF), mealworms (Tenebrio molitor), lesser mealworms (Alphitobius diaperinus), house cricket (Acheta domesticus, grasshoppers (Locusta migratoria), buffaloworms (Alphitobius diaperinus), cockroaches and domestic flies, whereby black soldier flies (Hermetia species, commonly called BSF), mealworms (Tenebrio molitor) and lesser mealworms (Alphitobius diaperinus) are more preferred.

For stabilizing insect meal an amount of at least one compound of formula (III) ranging from 10 to 1000 ppm, preferably ranging from 30 to 700 ppm, more preferably ranging from 100 to 500 ppm, based on the total amount of the insect meal, is usually sufficient.

Preferred embodiments of the present invention

The compounds of formula (III) with the preferences as given above are not only suitable for stabilizing fish meal, but also for stabilizing feed ingredients and feed. Preferences for feed ingredients and feed are given above and also apply here.

II. Feed

The compounds of formula (III) are not only suitable for stabilizing feed ingredients such as poultry meal, fish meal, insect meal and PUFA-containing oil, but also effective antioxidants for feed. Feed (or‘feedingstuff’) means any substance or product, including additives, whether processed, partially processed or unprocessed, intended to be used for oral feeding to animals. Feed in the context of the present invention is feed for aquatic animals and for terrestrial animals, as well as feed for insects.

For stabilizing feed an amount of at least one compound of formula (III) ranging from 10 to 500 ppm, preferably ranging from 30 to 300 ppm, more preferably ranging from

100 to 250 ppm, based on the total amount of the feed, is usually sufficient.

Further antioxidants

The compounds of formula (III) can be used in combination with one or more other antioxidants as described below.

In an embodiment of the present invention the feed of the present invention additionally comprises a mixture of 2-tert-butyl-4-methoxyphenol and 3-tert-butyl- 4-methoxyphenol, which is known under the name “BHA” (butylated hydroxyanisole).

In a further embodiment of the present invention the feed of the present invention additionally comprises ascorbyl palmitate. In another embodiment of the present invention the feed of the present invention additionally comprises BHA and ascorbyl palmitate.

Instead of ascorbyl palmitate other esters of ascorbic acid such as the esters of ascorbic acid with linear C12-20 alkanols, preferably the esters of ascorbic acid with linear C14-18 alkanols, may also be used, so that further embodiments of the present invention are directed to feed that additionally comprises esters of ascorbic acid with linear C12-20 alkanols, preferably esters of ascorbic acid with linear C14-18 alkanols, more preferably ascorbyl palmitate, whereby optionally BHA may also be present.

The feed may also comprise additionally alpha-tocopherol and/or gamma- tocopherol, whereby either an ester of ascorbic acid with a linear C12-20 alkanol with the preferences as given above or BHA or both may additionally be present. The feed itself is described in more detail below.

Feed for poultry

The feed for poultry differs from region to region. In the following Tables V and VI typical examples for diets in Europe and Latin America are given. These diets include cereals such as wheat, rye, maize/corn, minerals such as NaCl, vegetable oils such as soya oil, amino acids and proteins. Table V: European diet

Table VI: Latin American diet

Pet food

Pet foods are formulated to meet nutrient specifications using combinations of multiple ingredients to meet the targeted nutrient specification.

Poultry meal e.g. is an ingredient that is commonly found in Dog and Cat foods.

The nutrient specifications for a complete and balanced dog or cat food will meet or exceed the guidelines provided by AAFCO (American Association of Feed Control Officials). The ingredient composition of pet-food can include any legal feed ingredient so number of combinations are not quite infinite but close. Some examples of ingredient used in dog and cat foods can be found in Table VII below: Table VII:

For stabilizing pet food an amount of at least one compound of formula (I II) ranging from 10 to 500 ppm, preferably ranging from 30 to 300 ppm, more preferably ranging from 100 to 250 ppm, based on the total amount of the pet food, is usually sufficient.

Feed for fish

A typical example of feed for fish comprises the following ingredients, whereby all amounts are given in weight-%, based on the total weight of the feed for fish:

- Fish meal in an amount ranging from 5 to 15 weight-%, preferably fish meal in said amount comprising the compounds of formula (III) of the present invention;

- fish hydrolysates in an amount ranging from 0 to 5 weight-%;

- vegetable proteins in an amount ranging from 30 to 45 weight-%;

- binders, mainly starch, in an amount ranging from 9 to 12 weight-%;

- micro-ingredients such as vitamins, choline, minerals, mono calcium phosphate (“MCP”) and/or amino acids in an amount ranging from 3 to 6 weight-%;

- marine oil in an amount ranging from 5 to 10 weight-%, preferably marine oil in said amount comprising the compounds of formula (III) of the present invention; - vegetable oil in an amount ranging from 20 to 25 weight-%, preferably vegetable oil in said amount comprising the compounds of formula (III) of the present invention;

and whereby the amount of all ingredients sum up to 100 weight-%.

For stabilizing feed for fish an amount of at least one compound of formula (III) ranging from 10 to 1000 ppm, preferably ranging from 30 to 700 ppm, more preferably ranging from 100 to 500 ppm, based on the total amount of the feed for fish, is usually sufficient.

Process for the manufacture of compounds of formula (III)

Starting material (compounds of formulae (I) and (ID) and product (compound of formula (III))

“alkyl” and“alkoxy” in the context of the present invention encompass linear alkyl and branched alkyl, and linear alkoxy and branched alkoxy, respectively. Preferably R¹ and R³ are independently from each other H or methyl or ethyl, and R² is either H or methyl or ethyl or methoxy or ethoxy, preferably with the proviso that at least one of R¹, R² and R³ is H.

More preferably R¹ and R³ are independently from each other H or methyl, and R² is either H or methyl or methoxy, preferably with the proviso that at least one of R¹, R² and R³ is H.

The most preferred compounds used as starting materials and the most preferred compounds obtained as products by the process of the present invention are shown in Fig. 1 -3.

Fig. 1 shows the synthesis of 2,5,7,8-tetramethyl-2-(4-methylpentyl)chroman-6-ol (compound of formula (MI-1 )) starting from 2,3,5-trimethyl-1 ,4-hydroquinone (compound of formula (1-1 )) and 3,7-dimethyloct-1 -en-3-ol (compound of formula (11-1 )) .

Fig. 2 shows the synthesis of 2-methyl-2-(4-methylpentyl)chroman-6-ol (compound of formula (MI-2)) starting from 1 ,4-hydroquinone (compound of formula (I-2)) and 3,7-dimethyloct-1 -en-3-ol (compound of formula (11-1 )).

Fig. 3 shows the synthesis of 2-(4,8-dimethylnonyl)-2-methylchroman-6-ol

(compound of formula (MI-3)) starting from 1 ,4-hydroquinone (compound of formula (1-2)) and 3,7,1 1 -trimethyldodec-1 -en-3-ol (compound of formula (II-2)).

In a preferred embodiment of the present invention the molar ratio of the compound of formula (I) to the compound of formula (II) is in the range of 6.0:1 to 1 .1 :1 , preferably in the range of 4.0:1 to 1 .2:1 , more preferably in the range of 3.0:1 to 1 .3:1 , most preferably in the range of 2.0:1 to 1.5: 1.

In a further embodiment of the present invention all embodiments of the present invention with regard to the starting materials and the preferences as given above are realized.

Solvent mixture

In a preferred embodiment of the present invention the first of the two solvents is selected from ethylene carbonate, propylene carbonate, 1 ,2-butylene carbonate, and gamma-butyrolactone,

and the second of the two solvents is selected from hexane, cyclohexane, heptane, ortho-xylene, meta-xylene, para-xylene, mesitylene, pseudocumene, methyl tert- butyl ether, and toluene, whereby this preference is preferably combined with another preference, some other preferences or all preferences of the process according to the present invention .

In a more preferred embodiment of the present invention the first of the two solvents is ethylene carbonate or propylene carbonate, and the second of the two solvents is selected from either hexane, cyclohexane or heptane, preferably the first of the two solvents is ethylene carbonate and the second of the two solvents is heptane.

The term“hexane” encompasses n-hexane, as well as any mixture of the isomers of hexane. The same applies for heptane.

In a preferred embodiment of the present invention the volume ratio of the first solvent to the second solvent during the reaction is in the range of 1 :4 to 4: 1 , preferably the volume ratio of the first solvent to the second solvent is in the range of 1 :3 to 3:1 , more preferably the volume ratio of the first solvent to the second solvent is in the range of 1 :2 to 2: 1.

In another embodiment of the present invention the total amount of the two solvents is in the range of 1 to 10 kg, preferably in the range of 2 to 7 kg, more preferably in the range of 2.5 to 6 kg, per kg of the compound of formula (I).

In a further embodiment of the present invention all embodiments of the present invention with regard to the solvent and the preferences as given above are realized.

Acid catalyst

Examples of suitable acid catalysts are Bronsted acids and Lewis acids and any mixture thereof.

Examples of Bronsted acids are sulfuric acid, phosphoric acid, acidic ion-exchange resins (e.g. Amberlyst 15), acidic clays (e.g. Montmorillonite K-10), zeolites (e.g. HSZ-360), hydrochloric acid, trifluoroacetic acid, trichloroacetic acid, acetic acid, formic acid, methanesulfonic acid, benzenesulfonic acid, para-toluenesulfonic acid, ethanesulfonic acid, trifluoromethanesulfonic acid, bis(perfluoroalkyl- sulfonyl)methanes (R’S0₂)(R”S0₂)CH₂ wherein R’ and R” each signify independently from each other a perfluoroalkyl group of the formula C_nF_2n+i where n is an integer from 1 to 10, tris(perfluorosulfonyl)methanes (R’S0₂)(R”S0₂)(R”’S0₂)CH, wherein R’, R” and R’” each signify independently from each other a perfluoroalkyl group of the formula C_nF_2n+i where n is an integer from 1 to 10, and whereby at least two of R’, R” and R’” are identical perfluoroalkyl groups, or R’ signifies the

pentafluorophenyl group (-C₆F5) and R” and R’” each signify an identical perfluoroalkyl group of the above formula C_nF_2n+i, methanetrisulfonic acid, and bis(trifluormethylsulfonyl)imide, and any mixture thereof, whereby the use of single catalysts is preferred.

Examples of Lewis acids are Al(OTf)₃, Sc(OTf)3, Sc(NTf₂)₃, ScCl₃, Yb(OTf)₃, YbCl₃, Cu(OTf)₂, FeCl₂, Fe(OTf)₂, ZnCl₂, Zn(OTf)₂, Zn(NTf₂)₃, YCl₃, Y(OTf)₃, lnCl₃, lnBr₃, ln(OTf)₃, ln(NTf₂)₃, La(OTf)₃, Ce(OTf)₃, Sm(OTf)₃, Gd(OTf)₃, and Bi(OTf)₃ in the presence or absence of 2,2-bipyridine, and any mixture thereof, whereby the use of single catalysts is preferred.

Preferably the acid catalyst is para-toluenesulfonic acid, sulfuric acid,

methanesulfonic acid, Al(OTf)₃, Sc(OTf)₃, or ln(OTf)₃ and any mixture thereof, more preferably the acid catalyst is para-toluenesulfonic acid or Al(OTf)₃ or any mixture thereof. Hereby the use of single catalysts is preferred.

Thus, the more preferred acid catalyst is para-toluenesulfonic acid, sulfuric acid, methanesulfonic acid, Al(OTf)₃, Sc(OTf)₃, or ln(OTf)₃, and the most preferred acid catalyst is para-toluenesulfonic acid or Al(OTf)₃.

In a preferred embodiment of the present invention the amount of the acid catalyst is in the range of 0.001 to 5 mol equivalents, preferably in the range of 0.005 to 1 mol equivalents, more preferably in the range of 0.01 to 0.1 mol equivalents, relative to the amount of the compound of formula (II).

In the process according to the present invention the acid catalyst is recyclable which is a further advantage of the present invention.

In a further embodiment of the present invention all embodiments of the present invention with regard to the acid catalyst and the preferences as given above are realized. Reaction conditions

The reaction is preferably carried out at a temperature in the range of 70 to 160°C, more preferably in the range of 80 to 130°C, most preferably in the range of 90-105°C.

The reaction is preferably carried out at a pressure in the range of 0.8 to 20 bar (absolute), more preferably at a pressure in the range of 0.8 to 10 bar (absolute), most preferably at a pressure in the range of 0.8 to 5 bar (absolute).

In a further embodiment of the present invention all embodiments of the present invention with regard to the reaction conditions and the preferences as given above are realized. In the most preferred embodiments of the present invention all embodiments of the present invention with regard to the starting materials, the solvent, the catalyst and the reaction conditions including the preferences as given above are realized. The invention is now further illustrated in the following non-limiting examples.

Examples

Example 1 : Synthesis of 2.5.7.8-tetramethyl-2-(4-methylpentyl)chroman-6-ol (compound of formula (III-4) (see Fig. 1 )

A 1 .5 L 4-necked flask equipped with magnetic stirrer, oil bath, thermometer and argon supply 2,3, 5-trimethyl-1 ,4-hydroquinone (134 g, 853 mmol, 97%, 4.0 mol equiv.), was suspended in ethylene carbonate (300 ml.) and heptane (300 ml_), forming a 2-phase system upon warming to 60°C. Then, 3,7-dimethyloct-1 -en-3-ol (34.0 g, 213 mmol, 98%, 1 .0 mol equiv.) and para-toluenesulfonic acid

monohydrate (0.37 g, 2.13 mmol, 1 mol%) were added and the mixture was heated to reflux (85 °C internal temperature). After 60 min, the reaction mixture was cooled to 50 °C, diluted with water (500 ml.) and the phases were separated. The lower ethylene carbonate/water phase containing precipitated

trimethylhydroquinone was extracted with heptane (2x 250 ml_). The combined organic phases were extracted with water (500 ml_), dried over magnesium sulfate and concentrated in vacuo (45°C/50-20 mbar). The residue was purified by column chromatography, eluent gradient heptane/EtOAc 95:5 to 90:10 (w/w). The combined pure fractions were concentrated in vacuo (40°C/200-10 mbar) and dried under high vacuum at 40°C, furnishing 2,5,7,8-tetramethyl-2-(4- methylpentyl)chroman-6-ol (compound of formula (MI-4)) as colorless crystals (26.8 g g, 98.9% purity by quant. NMR, 91 mmol, 43% yield).

Example 2: Synthesis of 2-methyl-2-(4-methylpentyl)chroman-6-ol (see Fig. 2)

A 1500 ml. 4-necked flask with magnetic stirrer, oil bath, thermometer and argon supply was charged with 1 ,4-hydroquinone (95.0 g, 864 mmol, 99.8%, 4.0 mol equiv.), 3,7-dimethyloct-1 -en-3-ol (34.0 g, 216 mmol, 99.3 area% by GC, 1.0 mol equiv.) and dissolved in ethylene carbonate (400 ml.) and heptane (300 ml_), forming a 2-phase system. Then, para-toluenesulfonic acid monohydrate (0.38 g, 2.16 mmol, 98.5%, 1 mol%) was added and the mixture was heated to reflux. After 1 h, water (500 ml.) was added to the reaction mixture and the still warm reaction phases were separated. The lower ethylene carbonate phase was extracted heptane (2x 300 ml_). The combined heptane phases were dried over Na₂S0₄ and concentrated in vacuo (40° C/50-20 mbar). The residue was purified by column chromatography, eluent gradient heptane/EtOAc 95:5 to 85:15 (w/w). The combined pure fractions were concentrated in vacuo (40°C/200-10 mbar) and dried under high vacuum at 45°C, furnishing 2-methyl-2-(4-methylpentyl)chroman-6-ol as colorless oil (31 .7 g, 97% purity by quant. NMR, 124 mmol, 58% yield).

GC-HRMS: 95.4 area%. Calcd. for C₁₆H₂₄O₂ (M⁺) 248.1776, found 248.1800.

¹H NMR (300 MHz, CHLOROFORM-d) d 0.88 (d, J = 6.6 Hz, 6 H), 1 .12-1 .23 (m, 2 H), 1 .27 (s, 3 H), 1 .33-1 .46 (m, 2 H), 1 .46-1 .62 (m, 3 H), 1 .64-1 .89 (m, 2 H), 2.71 (t, J

= 6.9 Hz, 2 H), 4.51 (s, 1 H, OH), 6.54-6.63 (m, 2 H), 6.66 (d, J = 8.9 Hz, 1 H) ppm. ¹³C NMR (75 MHz, CHLOROFORM-d) d 21 .4 (1 C), 22.4 (1 C), 22.6 (2 C), 24.1 (1 C), 27.9 (1 C), 30.9 (1 C), 39.4 (1 C), 39.7 (1 C), 75.9 (1 C), 1 14.4 (1 C), 1 15.4 (1 C), 1 17.8 (1 C), 122.0 (1 C), 147.9 (1 C), 148.4 (1 C) ppm. Example 3: Synthesis of 2-(4.8-dimethylnonyl)-2-methylchroman-6-ol

(compound of formula (111-3)) (see Fig. 3) A 200 ml. 4-necked flask equipped with magnetic stirrer, oil bath, thermometer and argon supply was charged with hydroquinone (12.0 g, 109 mmol, 99.8%, 4.0 mol equiv.), 3,7, 1 1 -trimethyldodec-1 -en-3-ol (6.39 g, 27.2 mmol, 96.3 area% by GC, 1 .0 mol equiv.), and dissolved in ethylene carbonate (50 ml.) and heptane (50 ml.) forming a 2-phase system upon warming to 60° C. Then, para-toluenesulfonic acid monohydrate (0.10 g, 0.54 mmol, 2 mol%) was added and the mixture was heated to reflux. After 90 min, the reaction mixture was cooled to 80°C and the phases were separated. The lower ethylene carbonate phase was extracted with heptane (25 ml_). The combined organic phases were dried over sodium sulfate and concentrated in vacuo (40° C/50-20 mbar). The residue was purified by column chromatography, eluent gradient heptane/ EtOAc 95:5 to 85:15 (w/w). The combined pure fractions were concentrated in vacuo (40°C/200-10 mbar) and dried under high vacuum at 40 °C, furnishing 2-(4,8-dimethylnonyl)-2-methylchroman-6- ol (compound of formula (MI-3)) as light beige oil (4.95 g, 98.4% purity by quant. NMR, 15.3 mmol, 56% yield).

GC-HRMS: >99 area%. Calcd. for C₂₁H₃₄O₂ (M⁺) 318.2559, found 318.2370.

¹H NMR (300 MHz, CHLOROFORM-d) d 0.86 (d, J = ~ 6 Hz, 3 H), superimposed by 0.88 (d, J = 6.6 Hz, 6 H), 1 .04-1 .46 (m, 11 H), superimposed by 1 .27 (s, 3 H), 1 .46-1 .67 (m, 3 H), 1 .70-1 .87 (m, 2 H), 2.71 (t, J = 6.8 Hz, 2 H), 4.54 (br s, 1 H, OH), 6.53- 6.62 (m, 2 H), 6.66 (d, J = 8.5 Hz, 1 H) ppm.

¹³C NMR (75 MHz, CHLOROFORM-d) d 19.6, 21 .1 , 22.3, 22.6, 22.7, 24.1 , 24.8, 28.0, 30.8, 30.9, 32.7, 37.3, 37.5, 39.3, 39.78, 39.82, 76.0, 114.4, 1 15.4, 1 17.8, 122.0, 147.9, 148.4 ppm. Comparison example: reaction in one solvent only

2-meihyl-2-{4-methylpentyl)diroman-6-ol

yield: 20%

purity: S9%

“eq.” = mol equivalents; p-TsOH = para-toluenesulfonic acid, h = hour. Example 4: Antioxidant activities of compound of formulae (III-3) and (111-4) in fish oil and algal oil

The compounds of formulae (MI-3) and (111-4) have been tested. The blank oil, i.e. oil without any antioxidant, and oil containing“MNT” have been used as

benchmark. Any compound better in antioxidant activity than the blank oil indicates that it has antioxidant activity. The comparison with MNT gives an indication about the amount of the antioxidant effect, relative to the activity of MNT. “MNT” are mixed natural tocopherols commercially available as e.g.,“Tocomix 70 IP” from AOM (Buenos Aires, Argentina). Tocomix 70 IP comprises d-alpha- tocopherol, d-beta-tocopherol, d-gamma-tocopherol and d-delta-tocopherol, whereby the total amount of tocopherols is at least 70.0 weight-% and the amount of non-alpha tocopherols is at least 56.0 weight-%.

The compounds of formulae (111-3) and (MI-4) were evaluated primarily for their oxidative stability by the Oil Stability Index (OSI) measurements. Two different levels of these antioxidants (0.5 and 2 mg/g) were used in 5 g of natural fish oil (Product code: FG30TG) and used in the Oxidative Stability Instrument at 80°C with the air flow rate of 40 psi.

Crude algal oil (Lot# VY00010309) contained about 1 .6 mg/g of MNT prior to use in these experiments whereas fish oil did not contain any antioxidants. The compound of formula (MI-4) (see Table 2 below) and the compound of formula (MI-3) (see Table 3 below) were used at different times in the Oxidative Stability Instrument under similar operational conditions used for fish oil evaluations. MNT and the oil without any antioxidants were always used to compare. Also, the synergistic effect of only oil soluble compounds with ascorbyl palmitate was determined using the OSI values. The polymers generated at the end of the experiment were determined by LC (LC = liquid chromatography).

The solubility of the compounds used in the oxidative stability study are shown in Table 1 .

Table 1 : Solubility of compounds of formulae (MI-3) and (III-4) in fish oil

Oil Stability Indices (OSI) for these compounds at 500 and 2000 ppm levels, in comparison with the same amounts of MNT, are shown in Tables 2-3.

Table 2: Oxidative stability of FG30TG fish oil with compound of formula (III-4) (SD = standard deviation)

Table 3: Oxidative stability of FG30TG fish oil with compound of formula (MI-3) (SD = standard deviation)

The Protection Factors of the corresponding antioxidant compounds in fish oil are shown as a percentage in Tables 4-5.

The Protection Factors (PF) for each compound in oil were calculated in

percentage as:

100% x (OSI of the sample with compound - OSI of the sample without compound)

PF (%) =

OSI of the sample without compound

Table 4: Protection Factors of compound of formula (III-4) in FG30TG fish oil

Table 5: Protection Factors of compound of formula (MI-3) in FG30TG fish oil

Improvement of the oxidative stability of oil soluble compound of formula (111-4) when combined with AP is shown in Table 6 whereas Table 7 shows the same synergistic effect of compound of formula (MI-3) with AP. Table 6: Improvement of the effect of the compound of formula (111-4) in FG30TG fish oil using AP (SD = standard deviation)

Table 7: Improvement of the effect of compound of formula (MI-3) in FG30TG fish oil using a synergistic compound (AP) (SD = standard deviation)

Improvements of the Protection Factors of these oil soluble compounds with AP in fish oil are shown in Tables 8 and 9.

Table 8: Improvement of the Protection Factors of compound of formula (MI-4) with AP in FG30TG fish oil

Table 9: Improvement of the Protection Factor of compound of formula (111-3) with AP in FG30TG fish oil

Polymers generated at the end of the stabilization experiment of fish oil with compound of formula (MI-4) and AP are shown in Table 10.

Table 10: Reduction of polymers in FG30TG oil with a compound (AP) synergistic to compound of formula (III-4) (SD = standard deviation)

Tables 1 1 , 12 and 13 show the PV peroxide value), p- AV (g-anisidine value) and CD (conjugated dienoic acid %) of the fish oil samples stabilized with compounds of formulae (MI-3) and (MI-4), respectively.

Table 1 1 : Variation of PV (peroxide value) with compounds of formulae (MI-3) and (MI-4) in FG30TG

Table 12: Variation of p- AV (p-anisidine value) with compounds of formulae (MI-3) and (MI-4) in FG30TG

Table 13: Variation of CD (conjugated dienoic acid in %) with compounds of formulae (MI-3) and (III-4) in FG30TG

Results:

Compounds of formulae (II 1-3) and (111-4) showed very similar pattern of OSI in fish oil. The compound of formula (MI-4) had clearly lower OSI values than those of MNT in algal oil and seems to have possible prooxidant effect at higher levels (2 mg/g). The antioxidant effect of compounds of formulae (MI-3) and (MI-4) can be improved by combination with ascorbyl palmitate (“AP”) (Tables 6-7). Protection Factors of the compounds of formulae (MI-3) and (MI-4), including MNT, in fish oil could be improved by the addition of AP (Tables 8-9) indicating the possibility of combining AP with compounds of formulae (MI-3) and (MI-4) to improve the oxidative stability of matrices containing high amounts of unsaturated fatty acids such as marine oil. A combination of complex, polymeric compounds generated at the end of the oxidation cascade of unsaturated fatty acids indicate the levels of overall oxidation of the matrix. The generation of such polymers in fish oil containing these novel antioxidant compounds could be reduced considerably when AP was added as a synergistic compound (Table 10).

For the storage stability study oil soluble compounds were used in fish oil at 2 mg/g level only. Compared to the same level of MNT, all compounds showed much higher PVs than those of MNT (Table 1 1 ). There was no considerable variation in p-AV and CD (Tables 12-13) during the storage.

All compounds showed antioxidant properties in fish oil at different levels.

The oxidative stability of fish oil with compound of formula (III-4) is comparable to the antioxidative effect of MNT.

Claims

1 . A process for the manufacture of a compound of formula (III),

wherein n is 1 or 2, and

R¹ and R³ are independently from each other H or Ci-₅-alkyl, and R² is either H or Ci-₅-alkyl or Ci-₅-alkyloxy, with the proviso that at least one of R¹, R² and R³ is H, and wherein

the first of the two solvents is selected from ethylene carbonate, propylene carbonate, 1 ,2-butylene carbonate, gamma-butyrolactone and water, and the second of the two solvents is selected from hexane, cyclohexane, heptane, ortho-xylene, meta-xylene, para-xylene, mesitylene,

pseudocumene, methyl tert- butyl ether, and toluene.

2. The process according to claim 1 , wherein R¹ and R³ are independently from each other H or methyl or ethyl, and R² is either H or methyl or ethyl or methoxy or ethoxy, with the proviso that at least one of R¹, R² and R³ is H.

3. The process according to claim 1 , wherein R¹ and R³ are independently from each other H or methyl, and R² is either H or methyl or methoxy, with the proviso that at least one of R¹, R² and R³ is H.

4. The process according to any one or more of the preceding claims, wherein the first of the two solvents is ethylene carbonate or propylene carbonate, and the second of the two solvents is selected from either hexane, cyclohexane or heptane, preferably wherein the first of the two solvents is ethylene carbonate and the second of the two solvents is heptane.

5. The process according to any one or more of the preceding claims, wherein the volume ratio of the first solvent to the second solvent during the reaction is in the range of 1 :4 to 4:1 , preferably wherein the volume ratio of the first solvent to the second solvent is in the range of 1 :3 to 3: 1 most preferably wherein the volume ratio of the first solvent to the second solvent is in the range of 1 :2 to

2: 1 .

6. The process according to any one or more of the preceding claims, wherein the total amount of the two solvents is in the range of 1 to 10 kg, preferably in the range of 2 to 7 kg, more preferably in the range of 2.5 to 6 kg, per kg of the compound of formula (I).

7. The process according to any one or more of the preceding claims, wherein the acid catalyst is selected from Bronsted acids, Lewis acids and any mixtures thereof.

8. The process according to claim 7, wherein the acid catalyst is a Bronsted acid, and whereby the Bronsted acid is selected from sulfuric acid, phosphoric acid, acidic ion-exchange resins, acidic clays, zeolites, hydrochloric acid,

trifluoroacetic acid, trichloroacetic acid, acetic acid, formic acid,

methanesulfonic acid, benzenesulfonic acid, para-toluenesulfonic acid, ethanesulfonic acid, trifluoromethanesulfonic acid, bis(perfluoroalkylsulfonyl)- methanes (R’S0₂)(R”S0₂)CH₂ wherein R’ and R” each signify independently from each other a perfluoroalkyl group of the formula C_nF_2n+i where n is an integer from 1 to 10, tris(perfluorosulfonyl)methanes (R’S0₂)(R”S0₂)(R”’S0₂)CH, wherein R’, R” and R’” each signify independently from each other a perfluoroalkyl group of the formula C_nF_2n+i where n is an integer from 1 to 10, and whereby at least two of R’, R” and R’” are identical perfluoroalkyl groups, or R’ signifies the pentafluorophenyl group (-C₆F₅) and R” and R’” each signify an identical perfluoroalkyl group of the above formula C_nF_2n+i,

methanetrisulfonic acid, and bis(trifluormethylsulfonyl)imide.

9. The process according to claim 7, wherein the acid catalyst is a Lewis acid, and whereby the Lewis acid is selected from Al(OTf)₃, Sc(OTf)3, Sc(NTf₂)₃, ScCl₃, Yb(OTf)₃, YbCl₃, Cu(OTf)₂, FeCl₂, Fe(OTf)₂, ZnCl₂, Zn(OTf)₂, Zn(NTf₂)₃, YCl₃, Y(OTf)₃, lnCl₃, lnBr₃, ln(OTf)₃, ln(NTf₂)₃, La(OTf)₃, Ce(OTf)₃, Sm(OTf)₃, Gd(OTf)₃ and Bi(OTf)₃ in the presence or absence of 2,2-bipyridine.

10. The process according to any one or more of the claims 1 to 6, wherein the acid catalyst is para-toluenesulfonic acid, sulfuric acid, methanesulfonic acid, Al(OTf)₃, Sc(OTf)₃, or ln(OTf)₃, most preferably wherein the acid catalyst is para-toluenesulfonic acid or Al(OTf)₃.

1 1 . The process according to any one or more of the preceding claims, wherein the amount of the acid catalyst is in the range of 0.001 to 5 mol equivalents, preferably in the range of 0.005 to 1 mol equivalents, more preferably in the range of 0.01 to 0.1 mol equivalents, relative to the amount of the compound of formula (II).

12. The process according to any one or more of the preceding claims, wherein the molar ratio of the compound of formula (I) to the compound of formula (II) is in the range of 6.0:1 to 1.1 :1 , preferably in the range of 4.0:1 to 1 .2:1 , even more preferably in the range of 3.0:1 to 1.3:1 , most preferably in the range of 2.0:1 to 1 .5:1 .

13. The process according to any one or more of the preceding claims, wherein the reaction is carried out at a temperature in the range of 70 to 160°C, preferably in the range of 80 to 130°C, most preferably in the range of 90-105°C.

14. The process according to any one or more of the preceding claims, wherein the reaction is carried out at a pressure in the range of 0.8 to 20 bar (absolute), preferably at a pressure in the range of 0.8 to 10 bar (absolute), most preferably at a pressure in the range of 0.8 to 5 bar (absolute).

15. The process according to any one or more of the preceding claims, wherein the acid catalyst is recyclable.

16. 2-(4,8-dimethylnonyl)-2-methyl-chroman-6-ol.

17. Use of the compounds of formula (III) as antioxidants,

wherein n is 1 or 2, and

R¹ and R³ are independently from each other H or Ci-₅-alkyl, and

R² is either H or Ci-₅-alkyl or Ci-₅-alkyloxy, preferably with the proviso that at least one of R¹, R² and R³ is H.

18. The use according to claim 17, wherein the compound of formula (III) is the compound of formula (III-3) and/or the compound of formula (III-4). 19. The use according to claim 17 and/or 18, wherein the compounds of formula

(III) are used as antioxidants in feed or feed ingredients.

20. The use according to claim 19, wherein the feed is feed for aquatic animals, feed for terrestrial animals and feed for insects.

21 . The use according to claim 19, wherein the feed ingredients are selected from fish meal, poultry meal, insect meal, and PUFA-containing oils.

22. The use according to claim 21 , wherein the PUFA-containing oil is selected from marine oil, microbial oil, fungal oil, algal oil and PUFA-containing plant oil.

23. Feed for aquatic animals, feed for terrestrial animals and feed for insects containing at least one compound of formula (III),

wherein n is 1 or 2, and

R¹ and R³ are independently from each other H or Ci-₅-alkyl, and

24. Feed ingredients containing at least one compound of formula (III),

wherein n is 1 or 2, and

R¹ and R³ are independently from each other H or Ci-₅-alkyl, and