WO2012160442A1

WO2012160442A1 - Method to simultaneously enhance omega-3 and remove volatile contaminants

Info

Publication number: WO2012160442A1
Application number: PCT/IB2012/001086
Authority: WO
Inventors: Asgeir SAEBØ
Original assignee: Pharma Marine As
Priority date: 2011-05-20
Filing date: 2012-05-17
Publication date: 2012-11-29

Abstract

The present invention relates to processes for removing contaminants from oils and the oils produced thereby. In preferred embodiments, a distillation process for an oil suspected of containing contaminants comprises forming an endogenous volatile distillate fraction in said oil, and then distilling the oil to remove an endogenous volatile distillate fraction, wherein the distillate fraction is characterized by containing substantially less highly unsaturated fatty acids and substantially more environmental contaminants than the starting oil.

Description

Method to simultaneously enhance omega-3 and remove volatile contaminants

Cross-reference to Related Applications

This application claims the benefit of US Prov. Appl. 61/488,247 filed May 20, 2011, the contents of which are herein incorporated by reference in their entirety.

Field of the Invention

The present invention relates to processes for removing contaminants from oils and the oils produced thereby.

Background of the Invention

Molecular distillation of fats and oils can substantially reduce the content of volatile environmental substances contained in the source material. Adding a working fluid to the process can enhance removal of volatiles. The working fluid can either be added to the oil prior to distillation as outlined in 1 523 541 Bl or introduced into the condensed distillate flow as outlined in US Pat Application 2009/0118524 Al.

Production of omega-3 concentrates typically includes multiple production steps. The number of steps is increasing in response to demands of more and more purified oils in terms of organoleptic quality for inclusion in foods, stability towards oxidation, level of undesirable contaminants and finally concentration of the omega-3 fatty acids. Techniques to concentrate the omega-3 fatty acids are becoming more and more sophisticated to achieve as high level as possible. The number of production steps therefore tends to increase.

A triglyceride concentrate of omega-3 typically may for instance include the following 10 steps: Pre-purification or degumming of the oil, pesticide removal, alcoholysis to make ethyl esters, molecular distillation to pre-concentrate the omega-3 acids, urea-inclusion to remove saturated and monounsaturated fatty acids, enzymatic re-esterification to obtain triacyl glycerols, bleaching, molecular distillation to remove excess ethyl esters, deodorization and finally addition of antioxidants. Production of high purity products of single omega-3 fatty acids might even include another two or three steps.

For every step in the purification process, there is a potential danger of oxidation of the highly unsaturated fatty acids. Further, increasing number of production steps is undesirable from a quality control and quality assurance point of view. The high number of production steps also adds costs to the product and inevitably give a small loss of material in each step. From an environmental point of view, the energy footprint is also of concern when adding new production steps.

Summary of the Invention

In some embodiments, the present invention provides oil distillation processes for a source oil suspected of containing contaminants comprising: forming an endogenous volatile distillate fraction in the source oil, and distilling the source oil to remove the endogenous volatile distillate fraction, wherein the distillate fraction is characterized by containing substantially less highly unsaturated fatty acids and substantially more

environmental contaminants than the source oil prior to the distillation and the residual oil (the purified fraction remaining after distillation) contains substantially less environmental contaminants than the source oil and substantially more highly unsaturated fatty acids (such as DHA) than the source oil. In some embodiments, the residual oil contains a higher percentage of DHA and/or EPA weight/weight (w/w) than the source oil. In some particularly preferred embodiments, the residual oil contains a higher percentage of DHA weight/weight (w/w) than the source oil. In some preferred embodiments, the source oil is a marine oil. In some preferred embodiments, the residual oil contains less than about 2 pg/gram dioxin-like PCB^'s, preferably less than about 1 pg/gram dioxin-like PCB ^'s, more preferably less than about 0.5 pg/gram dioxin-like PCB ^'s, and most preferably less than about 0.1 pg/gram dioxin-like PCB ^'s.

In some embodiments, the endogenously produced distillate fraction comprises a free fatty acid, a monoglyceride, a methyl- or an ethyl ester or any combination thereof. In some embodiments, the highly unsaturated fatty acids are selected from the group consisting of eicosapentaenoic, docosahexaenoic, eicosapentaenoic, and arachidonic fatty acids and combinations thereof. In some embodiments, the process of forming the endogenous volatile distillate fraction comprises treating said oil with an enzyme selected from the group consisting of a lipase, phospholipase and combinations thereof. In some embodiments, the enzyme is non-specific as to position of the fatty acid on the triglyceride or phospholipid. In some embodiments, the enzyme hydrolyzes fatty acids other than EPA and DHA to a greater than EPA and DHA, especially DHA, i.e., the enzyme discriminates against hydrolysis of EPA and DHA, especially DHA. In some embodiments, the present invention provides an oil obtained by the processes described above, wherein said the comprises greater than 50% triglycerides, less than 5% monoglycerides, less than 2 % free fatty acids and/or ethyl esters, and less than 1 pg/g dioxin- like PCB^'s.

In some embodiments, the present invention provides an ethyl ester product obtained by ethanolysis and further refining the residual oil described above. In some embodiments, the present invention provides an ethyl ester or triglyceride concentrate produced from the ethyl ester product.

In some embodiments, the present invention provides a dietary supplement, oral dosage form, or functional food comprising an oil or concentrate as described above.

In some embodiments, the present invention provides an intermediate marine oil product comprising from about 70-99% free fatty acids, 1-30% monoglycerides and greater than 5 pg/g of dioxin like PCB^'s. In some embodiments, the present invention provides a free fatty acid intermediate oil containing greater than 500 pg/gram PCB congener 28 and greater than 2000 pg/g of PCB congener 52.

Description of the Figures

Figure 1 provides a flow diagram depicting the typical minimum processing steps to produce a distilled concentrate of omega-3 fatty acid ethyl esters.

Figure 2 provides a flow diagram of an exemplary process of the present invention.

Detailed Description of the Invention

The present invention provides a process by which a fraction of free fatty acids low in omega-3 fatty acids is developed in the oil by applying selected enzymes. Subsequent removal of the fraction by molecular distillation removes the free fatty acids and preferably yields an oil high in DHA. This process also removes environmental contaminants and makes a targeted contaminant removal distillation step superfluous. In some preferred embodiments, the present invention combines the process of removing environmental pesticides and non- omega-3 fatty acids into one operation. No working fluid needs to be added to the oil, and a concentrate of omega-3 fatty acids can readily be made.

In some preferred embodiments, use of non-position specific but fatty acid- discriminating lipases, coupled with molecular distillation enhances EPA, and particularly DHA content, and further removes most of the environmental contaminates in a single operation. This is in contrast to typical processing of omega-3 oils which requires a pesticide removal step in advance of the omega-3 concentrating steps.

Environmental contaminants are typically removed by molecular distillation since triacylglycerols are far less volatile than the contaminants. This technique has been used in the marine oil industry since about 1985 when molecular distillation was adopted by the company Broedr. Aarsaether in Alesund Norway and others. As another example, Acadia University on its website refers to a case study on Laer Products that also used this technology (Acadia Institute of Case Studies; on the world wide web at aics.acadiau.ca/case_studies/

laerproducts.html). The case study shows that before about 1991 the typical process included the following stages. The first stage removed volatile gases and preheated the oil. The second stage pulled the higher volatile components (such as aldehydes, free fatty acids, vitamins, cholesterol and PCBs) from the oil, leaving a purified oil product which was virtually odor free.

It subsequently was discovered that addition of a volatile fluid to the oil to be distilled improved removal of environmental contaminants. This system takes advantage of the very high volatility of many of the pollutants. As soon as the pollutants evaporated in the distiller and were flowing down the condensate surface along with a small amount of a volatile part of the oil, mainly free fatty acids, the pollutants started to evaporate from the condensate in the high vacuum plant. The minute quantity of oil in which the pollutants were condensed was simply so small that the concentration of the contaminants soon became oversaturated in the low atmosphere inside the distiller. The hot surface oil therefore was not optimally depleted of the environmental contaminants. Therefore, adding a volatile fluid to the oil or by other means secure that the volatiles distilled off could dissolve into something on the cool surface in the distiller improved the quality of the purified oil.

Breivik published a paper in 1997 on a method of producing a high concentrate of omega-3 in triglyceride form. The concentrated fatty acid ethyl esters were re-esterified to triglycerides by use of immobilized lipases under vacuum to remove the ethanol formed. Excess ethyl esters were distilled off by molecular distillation and in this process, the excess ethyl ester served as a working fluid to collect the environmental contaminants. The distillate (the working fluid), comprising 47.4% of the mixture, contained after distillation 0.03 mg/g of DDT and DDT metabolites and 0.005 mg/g a-HCH, 0.03 mg/g HCB, and 0.4 mg/g toxaphene whereas the target acyl glycerols did not contain detectable amounts of the pesticides. European Patent 1 523 541 B l (2009) teaches use of adding a working fluid to the oil to be purified and distilling off said working fluid. The content of working fluid added is preferably between 1% and 15% or more preferably between 3% and 8%.

The stripping technology in European Patent 1 523 541 Bl makes use of added ethyl esters as working fluids. This material is readily available as co-stream products from the concentration of omega-3 fatty acids in ethyl ester form. The omega-3 depleted fractions of less volatility than the longer chain omega-3 fatty acid ethyl esters are added to the oil to be purified. After collection and when enriched in environmental contaminants they are used as biodiesel in the factory.

As mentioned above, the working fluid is added because the contaminant to be distilled off needs a fluid to be dissolved in, facilitating removal of contaminants in the low pressure atmosphere of the distiller. The dissolving fluid does not need to be added in front of the distillation, but may alternatively be introduced into the condensing area in the distiller as outlined in US Pat Application 2009/0118524 Al. Here, the evaporated volatile components are brought into contact with a washing liquid introduced to the system.

In Figure 1, the typical minimum processing steps to produce a distilled concentrate of omega-3 fatty acid ethyl esters are depicted in a flow diagram. The process includes the following steps: Alkali wash, pesticide removal, alcoholysis to yield fatty acid ethyl esters, distillation to enhance omega-3 esters, cold filtration to remove high melting moieties, bleaching to remove trace metals and prooxidants, and finally addition of antioxidants. As outlined above, removal of environmental contaminants is greatly improved by adding a working fluid. The current state of the art is either to add an ethyl ester to the oil prior to distilling or to introduce the fluid into the distillate flow in the distiller.

The present invention omits addition of working or washing fluids and also the initial alkali wash to remove free fatty acids. The technology is particularly efficient in the production of concentrates of docosahexaenoic acid (DHA), but is also applicable for concentrates of eicosapentaenoic acid (EPA), particularly as new and selective enzymes become available that preferentially hydrolyze other fatty acids than EPA. Enzymes particularly suitable for enclosed technology include but are not limited to enzymes obtained from Candida rugosa, previously Candida cylindracaea.

Y.Tanaka et. al. (1992) reported that a lipase from Candida cylindracaea was capable of enhancing the content of DHA in the glyceride fraction in fish oil from 8.9% to about 30%. They also reported that the enzyme worked well on tuna oil. Likewise, the lipase was also shown to perform well on Brazilian Sardine Oil, enhancing DHA from 10.2% to 22.5% in the glyceride fraction. (Carvalho et. al. 2002).

Farmed salmon oil has also been the object of enzymatic hydrolysis experiments. Enhancement (or reduction) of EPA and DHA in the glyceride fraction was reported for enzymes obtained from Rhizomucor miehei, Rhizopus delemar, Rhizopus oryzae,

Burkholderia cepacia and Candida rugosa (previously C. cylindracaea). Lipases from Burkholderia cepacia and Candida rugosa modestly enhanced EPA in the glyceride fraction, the first one at the expense of DHA. Both Rhizopus oryzae and Candida rugosa enhanced markedly DHA (from 6.93 to 11.14 and 19.39% respectively) and both also slightly enhanced EPA (D. Kahveci et. al. 2010).

Candida rugosa is a non-position specific enzyme but apparently discriminates against hydrolysis of EPA and particularly strongly against DHA. EPA and particularly DHA is typically a sn^'2- moiety in the triglycerides of fish and squid oils. Therefore, Ί 3, -specific enzymes have been tested as tools for enhancement of EPA and DHA in the glyceride fraction. A Mucor miehei preparation has been shown to be a valuable tool for preparation of monoacylglycerols of EPA and DHA in high purity (S.Nieto et. al. 1999). However, this work did not disclose relative composition of the glyceride fraction, but rather how a very pure fraction was obtained after subsequent clean up steps.

However, even if a Ί 3 specific lipase was able to produce a monoglyceride fraction of very high purity of DHA, such a blend would not be suitable for the disclosed technology since most of the monoglycerides would be distilled off along with the environmental contaminants and the free fatty acids. For example, if a triacyl glycerol contained oleic acid moieties in position Ί and ^'3 and DHA in position ^'2, a Ί-^'3 position specific enzyme could easily produce high yields of DHA monoglycerides and oleic acid in free form. However, the ^'2-DHA monoglyceride would be distilled off together with the oleic acid at the temperature required to obtain a residual oil substantially depleted in free fatty acids. The content of free fatty acids should preferably be well below 1% to be suitable for further refining. In preferred embodiments of the present invention, non-position specific lipases, preferably discriminating strongly against EPA and/or DHA, are utilized as a catalyst to develop the free fatty acids to be removed when concentrating EPA/DHA.

The process of the present invention comprises the following basic steps: Enzymatic treatment, pre-concentration by distillation, alcoholysis, purifying distillation, cold filtration, bleaching and final antioxidant addition. A flow scheme is provided in Figure 2. In the initial enzymatic treatment of the crude oil, up to 50% of the oil, preferentially about 30-40% is hydrolyzed into free fatty acids. The free fatty acid fraction preferably comprises fatty acids that not of interest for retention in the final product. In oils particularly high in DHA and low in EPA, up to 50% of the oil may preferentially be hydrolyzed. In the following step, the free fatty acid fraction of the oil is distilled off. The removal of this large amount of free fatty acids involves a two step removal procedure and use of high vacuum and relatively high temperature. The high content of free fatty acids is challenging to the vacuum system and the design of the plant. Free fatty acids tend to develop volatile moieties that can counteract the ultra vacuum typically achievable for ethyl esters. Further hydrolysis of mainly saturated and monounsaturated fatty acids yields a free fatty acid blend with a high melting point. Nevertheless, it has been found that a targeted plant design and process including a two stage plant are capable of removing the low omega-3 free fatty acid fraction down to a residual content of about 0.5%.

The removal of the low omega-3 fraction also removes contaminants such as environmental pesticides down to a desired level, making a specific targeted pesticide removal step superfluous. In some embodiments, the oil obtained after distillation is converted into ethyl esters by alcoholysis and further distilled, either to purify the oil, or optionally, to further concentrate the omega-3 fatty acids. Purification involves running the ethyl ester through the distiller to collect the product as a distillate and removing the residual part containing non- esterifed glycerides, cholesterol, soap, pigments and other high boiling point impurities. From a raw material containing about 20% DHA, a DHA concentrate of ethyl esters containing about 40% DHA is achievable without any conventional ethyl ester fractionation. However, in some embodiments of the invention, a further concentration can be performed by repeated distillations on the ethyl ester fraction.

In still other embodiments, additional techniques can be employed to further enhance the content of EPA and or DHA. These techniques include but are not limited to urea inclusion (complexation), supercritical chromatography, and distillations in combination with use of enzymes.

The distilling steps of the present invention have a substantial, positive impact on energy footprint and plant throughput. First, conventional techniques teach addition of a working fluid to the oil to be purified. The processes of the present invention do not add any working fluid, but typically produce in the oil a low omega-3 fraction that will function also as a working fluid when removed to enhance EPA and or DHA. Second, an advantage in terms of energy footprint of the processes of the present invention is that 40 to 50% or even more of the oil is removed before the purifying process proceeds with cleaning-up steps. This results in a 40-50% reduction in use of ethanol, catalyst, and energy in the alcoholysis step. Furthermore, the quantity of oil passed through the distillation for a further concentration is substantially reduced.

Free fatty acids are far from desirable to add as a working fluid due to vacuum problems in the distiller, solidifying problems in the lines (high melting point) and gasses in the atmosphere (working environment) calling for use of protection devices or properly systems. Despite these difficulties, the disclosed technology greatly improved the production capacity and reduced the energy footprint of the plant.

The maximum limit for Dioxin-like PCB^'s in food oils in the European Union is 3 pg/gram. Dioxin-like PCBs have "dioxin-like" properties. After distillation according to the present invention, the residual product oil contains less than about 2 pg/gram dioxin-like PCB ^'s, preferably less than about 1 pg/gram dioxin-like PCB ^'s, more preferably less than about 0.5 pg/gram dioxin-like PCB^'s, and most preferably less than about 0.1 pg/gram dioxin- like PCB^'s.

EXAMPLES

Example 1:

3.3 grams of a commercial powder of Candida rugosa enzyme was dissolved in 338 gram of water and mixed with 550 grams of crude oil from saithe. The batch was stirred for 3 hours and 20 minutes at 40 C before stirring was stopped and layers allowed to separate. Bottom enzyme water was drained off and 1000 ml of fresh water added to the batch followed by stirring and heating to 65 C to neutralize the enzymes. Oil was separated after standstill and dried under vacuum. A sample of the oil was separated by HPLC in a

triglyceride/diglyceride (TG/DG) fraction and a free fatty acid fraction (FFA). The TG/DG fraction contained 10.53% EPA and 29.15% DHA whereas as FFA fraction contained 4.94% EPA and 4.86% DHA. The residual omega-3 concentrate will after removal of free fatty acids (60%) by molecular distillation contain very low levels of environmental contaminants. Example 2:

A full scale test was performed with Candida rugosa lipase on Squid oil. 36000 kg of oil with a free fatty acid content of 7.6% was subjected to enzymatic hydrolysis with a lipase dissolved in water and added to the oil and stirred for 3 hours at 38 C. The free fatty acid content was enhanced to 35.1%. The oil was then distilled to remove the free fatty acids and the residual oil contained 0.8% free fatty acids. The DHA content of the oil was 35.3%, up from 26.8%.

Example 3:

In a full scale experiment 1800 kg of oil was filled into a reactor. Then 1 kg of Lipase

OF was dissolved in 600 liter of water and added to the oil while stirring. The oil contained before addition of enzymes 9.97 % of free fatty acids. After 29 hours, stirring was stopped and bottom layer with water pha O_iflt aer remo^se with enzymes was drained off. Another

Table 2. PCDD/F congeners pg/gram in the distillates and the residual oil.

Table 3. Tox e ivalents PCDD/F / ram

Table 8. Toxa hene Parlar Con eners mg/k

Table 9. Or anochloride Pesticide, P rethriodes, PCBs m /k

*total DDT

Claims

Claim We claim:

1. An oil distillation process for a source oil suspected of containing contaminants

comprising:

forming an endogenous volatile distillate fraction in said source oil, and distilling said source oil to remove said endogenous volatile distillate fraction, wherein said distillate fraction is characterized by containing substantially less highly unsaturated fatty acids and substantially more environmental contaminants than said oil prior to said distillation.

2. Process according to claim 1, wherein the source oil is a marine oil.

3. Process according to claim 1 or 2, wherein the endogenously produced distillate

fraction comprises a free fatty acid, a monoglyceride, a methyl- or an ethyl ester or any combination thereof.

4. Process according to any of claims 1 to 3, in which said unsaturated fatty acids are selected from the group consisting of eicosapentaenoic,- docosahexaenoic, eicosapentaenoic, and arachidonic fatty acids and combinations thereof.

5. Process according to any of claims 1 to 4, wherein the process of forming said

endogenous volatile distillate fraction comprises treating said oil with an enzyme selected from the group consisting of a lipase, phospholipase and combinations thereof.

6. An oil obtained according to any of claims 1 to 5, wherein said oil comprises greater than 50% triglycerides, less than 5% monoglycerides, less than 2 % free fatty acids and/or ethyl esters, and less than 1 pg/g dioxin-like PCB^'s.

7. An ethyl ester product obtained by ethanolysis and further refining of said oil of claim 6.

8. An ethyl ester or triglyceride concentrate produced from said ethyl ester of claim 7.

9. A dietary supplement, oral dosage form, or functional food comprising an oil or concentrate according to any of claims 6 to 8.

10. An intermediate marine oil product comprising from about 70-99% free fatty acids, 1-

30% monoglycerides and greater than 5 pg/g of dioxin like PCB ^'s.

11. A free fatty acid intermediate oil containing greater than 500 pg/gram PCB congener 28 and greater than 2000 pg/g of PCB congener 52.