BR-112020022516-B1 - PROCESS FOR ENZYMATIC DEGUMMING OF OIL

Abstract

PROCESS FOR ENZYMATIC DEGUMMING OF OIL. The present invention relates to a process for reducing the amount of intact phospholipids in a triacylglyceride oil, which comprises incubating the oil with a polypeptide that has phospholipase A1 activity, wherein the polypeptide comprises a polypeptide that has at least 80% identity with the mature amino acid sequence of SEQ ID NO: 1.

Inventors

• YING ZHA
• Arjen Sein
• Willem Bijleveld

Assignees

• DSM IP ASSETS B.V

Dates

Publication Date

20260317

Application Date

20190506

Priority Date

20180507

Claims (10)

1. 1. Process for reducing the amount of intact phospholipids in a triacylglyceride oil, characterized by comprising incubating the oil with a polypeptide that has phospholipase A1 activity, wherein the polypeptide comprises a polypeptide consisting of the mature amino acid sequence of SEQ ID NO: 1, and wherein at least 85% of the amount of intact phospholipids is reduced and the polypeptide is capable of reducing at least 85% of the amount of intact phospholipids originally present in the oil when phospholipase A1 is incubated with the oil in an amount of 0.28 mg of active protein/kg of oil at a temperature of 55°C, 60°C, 65°C and/or 70°C for 4 h.
2. 2. Process according to claim 1, characterized in that the phospholipids comprise phosphatidic acid, phosphatidyl ethanolamine, phosphatidylinositol and/or phosphatidylcholine.
3. 3. Process according to claim 1 or 2, characterized by further comprising a step of adding an acid to the oil.
4. 4. A process according to any one of claims 1 to 3, characterized by further comprising a step of adding water to the oil.
5. 5. A process according to any one of claims 1 to 4, characterized by further comprising a step of adding a caustic to the oil.
6. 6. Process, according to any one of claims 3 to 5, characterized in that the step of adding acid, water and/or caustic is carried out before incubating the oil with the phospholipase.
7. 7. A process according to any one of claims 1 to 6, characterized by further comprising incubating the oil with a polypeptide having phospholipase C activity, a polypeptide having phosphatidylinositol phospholipase C activity and/or a polypeptide having phospholipase A2 activity.
8. 8. A process according to any one of claims 1 to 7, characterized by further comprising separating phosphorus-containing components from oil.
9. 9. A process according to any one of claims 1 to 8, characterized in that the oil comprises crude oil or oil degummed with water.
10. 10. A process according to any one of claims 1 to 9, characterized in that the oil comprises a vegetable oil, algae oil, animal oil or fish oil.

Description

Field [0001] The present invention relates to a process for reducing the amount of phospholipids in a triacylglyceride oil using an enzyme that has phospholipase A1 activity. Background [0002] Crude vegetable oils obtained from pressing or solvent extraction methods are a complex mixture of triacylglycerols, phospholipids, sterols, tocopherols, free fatty acids, trace metals, and other secondary compounds. In soybean oil processing, soybean seeds may first be flaked before hexane extraction to obtain flake oil. In another commonly known process, the seed is first treated with an expander before extraction, resulting in expander oil. The latter generally results in a higher oil yield, but also a higher phospholipid content. During the preparation of other oils, such as canola or rapeseed oil, the seeds are first pressed, resulting in a pressed oil fraction. The press cake may be further treated with a solvent to produce an extracted oil fraction, and the two fractions combined are known as crude canola, rapeseed, or sunflower oil. [0003] It is desirable to remove phospholipids, free fatty acids, and trace metals to produce a high-quality edible oil. The most commonly used processes in the industry are aqueous or wet degumming, acid degumming, caustic refining, and enzymatic degumming or refining. In general, the removal of phospholipids generates most of the losses associated with the degumming of vegetable oils. Since most phospholipid molecules possess both a hydrophilic functional group and a lipophilic chemical portion consisting of a glycerol with two fatty acid chains, they tend to be excellent natural emulsifiers. Therefore, it is desirable to hydrolyze phospholipids into their lyso- or phosphorus (glycerophosphate) forms and then reduce the emulsifying property. The main phospholipids in vegetable oils are phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylinositol (PI), and phosphatidic acid (PA). [0004] Several processes are known for enzymatic degumming or enzymatic refining of vegetable oils, using enzymes with phospholipase activity, such as phospholipase A1, phospholipase A2, phospholipase C or phosphatidylinositol phospholipase C. [0005] Document WO9705219 discloses a process for reducing the content of phosphorus-containing components in vegetable oils using a mixture of phospholipase enzymes from Aspergillus niger, comprising a phospholipase A2 and/or phospholipase A1 activity and a lysophospholipase activity. [0006] Document EP0575133 teaches about a phospholipase A1 (PLA1) enzyme from an Aspergillus oryzae strain and an Aspergillus niger strain and about the use of these phospholipases to prepare lysophospholipids from a phospholipid substrate, for example, derived from animals, plants and microorganisms. Document EP0575133 reveals that the residual activity of A. niger PLA1, after heat treatment at 70 °C, was only about thirty percent (30%) of the residual activity after heat treatment at 50 °C and 60 °C. A. oryzae PLA1 did not exhibit any activity after heat treatment at 70 °C. Document EP0575133 does not teach a process for enzymatic degumming of an edible oil. [0007] Document US20160289658 discloses a phospholipase derivative from Talaromyces leycettanus. This phospholipase showed relatively high thermostability and demonstrated greater activity at 70 °C than the commercial enzyme preparation Lecitase Ultra. [0008] Document WO2011/051322 discloses a phospholipase from Aspergillus fumigatus that hydrolyzed phospholipids in soybean oil at temperatures of 55 °C and 60 °C. [0009] There is a need for an improved process to reduce the phospholipid content in a tricalcylglyceride oil using phospholipases that are active over a wide temperature range. Summary [0010] The present invention relates to a process for reducing the amount of intact phospholipids in a triacylglyceride oil, which comprises incubating the oil with a polypeptide that has phospholipase A1 activity, wherein the phospholipase A1 comprises a polypeptide that has at least 80% identity with the mature amino acid sequence of SEQ ID NO: 1. It was found that the phospholipase A1 that has at least 80% identity with the mature amino acid of SEQ ID NO: 1 was able to reduce at least 85% of the phospholipids originally present in a triacylglyceride oil over a wide temperature range of 55 °C to 70 °C in 4 h. A phospholipase A1 in a process as disclosed herein is a phospholipase A1 that hydrolyzes at least 85%, 86%, 87%, 88%, 89%, or at least 90% of the amount of intact phospholipids originally present in a triacylglyceride oil when incubated with the oil in an amount of 0.28 mg of active protein/kg of oil at a temperature between 55°C, 60°C, 65°C, and 70°C for 4 h. Definitions [0011] A “mature polypeptide” is defined in this document as a polypeptide in its final form and is obtained after translation of an mRNA into a polypeptide and post-translational modifications of said polypeptide. Post-translational modificatio