Processing of Crude Algal Oils and Production of EPA and DHA Concentrates

Rick Green, Ph.D.

POS Bio-Sciences

Saskatoon, SK

BIO Pacific Rim Summit

December 8, 2014

Overview of POS Bio-Sciences

• POS is a fee-for-service technology

development organization.

• Specialize in process development for

ingredients along with globally recognized

analytical services.

• Extraction and isolation of components from

plant and microbial sources.

• Extensive experience in algal oil processing

and fractionation to produce EPA and DHA

concentrates.

• Manage a multi-scale processing facility.

• 38 year technology development history.

Overview of POS Bio-Sciences

New process/product concept

Laboratory

(2 kg)

Mini-pilot scale trials

(20 kg)

Pilot scale development

(200 kg - Tonnes)

Custom manufacturing

(10 T – 600 T)

Commercial production

Components of Extracted

Microalgae Crude Oil

Lipids*

• Neutral

• Triacylglycerols (TAG)

• Unsaturated (including n-3)

• Saturated

• Hydrocarbons

• Sterols

• Waxes

• Pigments – chlorophyll, astaxanthin

• Tocopherols

• Polar

• Phospholipids

• Glycolipids

• Free fatty acids

Elements

• S, Cl, Na, heavy metals

* Some components

beneficial, some not.

Non-lipid components

• Undesirable Contaminants

• Oxidized compounds

• Pesticide residues

• Proteins/peptides

• Polysaccharides

• Sugars

• Moisture

• Further processing of crude oils is required to remove the

objectionable components – can have significant effect on yields.

Components of Extracted

Microalgae Crude Oil

Purification and Fractionation of

Microalgae Oil

• Increase stability of the oil

• Purify for further processing

• Increase functionality of the oil

• Produce valuable co-products

• Enhance economic viability

Co-products

• Generally, co-products for functional foods,

nutrition and health benefits are higher in

value

• E.g. astaxanthin, EPA, DHA

Downstream Processing Flow for Oil and

Co-products from Microalgae

Microalgae

Concentrated

unsaturated fraction

(DHA, EPA)

Crude Oil

Refined oil,

Saturates

Meal

feed

food ing.

fertilizer

protein

carbohydrates

pigments,

antioxidants,

antimicrobials

(Should be processed

under heavy N2)

Crude oil

Degumming

Alkali-refining

BleachingDeodorization

AO fortification &

Packaging

Water washing

Fractionation

Processed oil

Omega-3 rich

fraction

Waxes, polar cmpds.

Phospholipids & Glycolipids

Free Fatty acids (as soaps)

Pigments,

elements

n-3 Oil

Microalgal Oil Processing Schematic

Wash or Trisyl Treatment Trace soaps

Water washing

• Crude oil with 15% (w/w) warm water (~55-60°C under N2).

• Centrifuge (at ~55-60°C) – separation of water and oil phases.

Target:

To remove – Sugars

Polysaccharides

Fines

Mucilaginous compounds

Proteins, etc.

Avoid if oil is high in polyunsaturates

Water Washing

Crude Algal Oil

Desludger

Hydratable gums – water degumming

Non-hydratable gums – acid degumming

• Oil is mixed with water (2% w/w) or acid (0.2% w/w) & water (2% w/w)

Acid: Citric or Phosphoric acid.

• Centrifuge: gives semi-solid gum phase and liquid oil phase

(degummed oil).

Washed Oil

Degumming

• First step of the refining process, designed to remove

phosphatides (gums) that interfere with subsequent processing.

Alternative to Degumming

• Mixing the oil in acetone will cause precipitation of the polar lipids

(phospholipids and glycolipids)

• Need to use explosion proof equipment and a solvent rated facility

• A more expensive method

Precipitation of Polar Lipids in Acetone

Degummed oil

• Involves the removal of free fatty acids (FFA)

• Sufficient caustic is added to convert the FFA to soaps (FFA salts) –

these are water insoluble.

• Calculate the required amount of caustic (dilute NaOH solution or

Baumé) based on free fatty acid content.

• Required amount of caustic mixes with degummed oil at 60C for

minimum of 30 min.

• Centrifuge at 70°C to separate soap and liquid-oil phase

(alkali-refined oil).

Alkali Refining

Continuous

Refining Separator

Alkali RefiningFFA Soaps

OH-C-R + KOH

O

K+-O-C-R + H2O

O

Alkali-refined oil

• Trisyl - an amorphous, synthetic silica hydrogel used to remove small

amounts of soaps formed during alkali refining of the oil.

(in lieu of water washing)

• Due to the polar nature of Omega-3 containing Algae oil, water washing

is not suitable for removing soap (yield losses may be very high).

• Calculate the required amount of Trisyl, based on the remaining soap.

• Trisyl mix with refined oil at 60°C for ~30 min.

• Filter using frame filters to separate soap adsorbed Trisyl-solids

and Trisyl-treated oil.

Trisyl Treatment

Bleaching

• Oil is mixed with an adsorptive clay to remove pigments

(e.g. chlorophyll) and elements/metals.

• Also removes some breakdown products of any fatty

acid/TAG oxidation (e.g. peroxides, ketones)

Filter PressReactor for bleaching

Trisyl treated oil

Deodorization

Continuous Column Deodorizer

(250 kg/hr)

Column packing

Remove odors and flavors from high oils/fractions.

Bleached oil

Fractionation of Oil & Production of

DHA & EPA Concentrates

Dr. Udaya Wanasundara

Principal Scientist

Lipids & Co-Products

Low temperature crystallization (in solvent)

Urea Adduct Formation

Short Path Distillation/Fractional Distillation

Supercritical fluid extraction & fractionation

Chromatographic separation (CPC, HPLC)

Esterification is usually required prior to the concentration

All these methods have advantages and disadvantages

Important consideration should be product quality and the cost of production

Methods for Concentration of Omega-3 Fatty Acids

Fatty Acid Melting point (C)

Melting point of a FA depends

on the chain length and the

number of double bonds

(unsaturation) present.

Saturates – high MP

PUFA – low MP

Low Temperature Crystallization

Example Conditions

• Algae oil (TAG or FFA)

• Solvents: Hexanes or Acetone

• Oil-to-solvent ratio: 1:4 (w/v)

• Temperature: -25oC

• Time: 24h with slow mixing

• Separate crystals by filtration

• Evaporate solvent from liquid

fraction (non-crystallized

fraction)

Low Temperature Crystallization

Non-crystallized fraction

(PUFAs)

Crystallized fraction

(Saturated fatty acids)

Jacketed Tank

Enrichment of PUFAs by Low Temperature Crystallization

AcetoneHexanes

12.0

15.1

37.2

TAG

8.2

10.1

27.8

TAG

13.8

17.5

40.6

11.8

12.5

31.6

6.8

8.4

22.2

EPA (%)

DHA (%)

Total 3 (%)

FFAFFA

Starting

Marine

Oil

Fatty Acid

Esters

Urea Adduct Formation

5.67Å

CO

HN2 HN2

FFA/EE

mixture

Urea:CO(NH2)2

(Tetragonal)

Urea & Straight Chain

Molecules (Hexagonal)

PUFA

(NUCF)+

UCF

8-12ÅFA

• In a FFA or ethyl ester

(EE) mixture, saturates

and mono-unsaturates

are complexed with urea

(UCF).

• Presence of double

bonds increases the

bulk of the molecule and

reduces the likelihood of

its complexation with

urea, i.e. PUFAs remain

in the liquid and are

referred to as non-urea

complexing fraction

(NUCF).

• Dissolve urea in 95% ethanol or methanol.

• Add either FFA or Ethyl ester form of Omega-3 rich oil

• Mix and leave at room temperature or ~4C to form crystals (Urea/fatty acid complex or adduct).

During crystallization, straight saturated and mono-unsaturated fatty acids are form complex with urea (Urea Complexed Fraction, UCF) and PUFAs remain in the liquid (Non-Urea Complexed Fraction, NUCF).

• Solid UCF (with saturates and mono-unsaturates ) and liquid NUCF (with PUFAs)

can be separated by filtration.

Urea Adduct Formation

Enrichment of EPA- and DHA-EE by Urea Method

6.8 4.2 8.4

10.9 2.4 67.6

3:1

57.1

98.8

4:1

Starting oil

After urea

Urea-to-EE Ratio(w/w)

Marine

EPA DPA DHA

Algal

DHA

Values are in w/w%

Short Path Distillation

• Uses the differences in boiling point and

molecular weight of fatty acid esters

• Saturated short chain acid-esters – lower

boiling point and more volatile than long

chain high MW fatty acid-esters, such as

PUFA.

• Distillation of fatty acid esters is carried out

under vacuum (0.1-4.0 mmHg)

• Distillation temperatures (150-210oC).

Fractionation

Short Path Distillation

• Fractionation of oils, waxes• Preparation of value-added fractions (e.g. Omega-3 rich concentrates)

Short Path Distillation of 3 Fatty Acid-Ethyl Esters

11.8

12.2

26.2

53.1

23

11.8

7.0

15.5

38.2

50

7.0

4.2

8.8

23.2

EPA-EE

DPA-EE

DHA-EE

Total 3-EE

Yield (%)

Trial-2Trial-1Starting Algal Oil Esters Fatty Acid Esters

(%)

Conditions Trial-1 Trial-2

Temperature (oC) 170 190

Pressure (mm Hg) 1.5 1.5

Holding Time (min) 7 7

Summary of Scale-up Processing for Microalgal Oil

• Increase purity by degumming, alkali refining, Trisyl

treatment, bleaching and optionally deodorizing.

• Prepare DHA and EPA concentrates by fractionation of

the refined oil.

• Viable large scale fractionation techniques include low

temperature crystallization, urea adduct formation and

fractional distillation (short path distillation).

Thank You!POS Bio-Sciences118 Veterinary RoadSaskatoon, SKCANADA S7N 2R4

Tel. (306) 978 2800

email: rgreen@pos.ca