Novel Separation Techniques for Isolation of Fatty Acids and Oil By-Products

National Conference on

“Newer Oleo-chemicals: Production & Industrial Application”

Organised By

Centre of Excellence on

‘Applied Research, Training & Education in Lipid Science’Department of Oil & Paint Technology,

Harcourt Butler Technological Institute, KanpurJanuary, 10-11, 2015

Sadanand Patel & Sanjay Kr Singh

Department of Oil & Paint Technology

HBTI, Kanpur

sanjayhbti19@gmail.com

Contents

Conclusion

By products

Enzymatic & Urea Complexion

Counter Current Chromatography

Need & Fundamentals

Need???

Potential health benefits of bioactive compounds

Challenge of refineries

Profitability of processing operations

.

.

Separation

Density

Boiling Point

Melting Point

Centrifugal

Separation

Distillation

Crystallization

Principle of Separations

Molecular Properties

Bulk Properties

Melting Point Boiling Point Density

Position & Nature of Bonds Packing Structure Molecular Dimensions

Liquid Chromatography

Packing Materials Type

Adsorption

Ion Exchange

Gel Filtration

Non Packing Materials Type

Centrifugal Field

Gravitational

Preparative Liquid Chromatography

Counter Current Chromatography

This technique includes:

Liquid-liquid partition,

Countercurrent distribution of solute mixture

between two immiscible liquid phases,

In the most distinct variants of CPC, one liquid phase remains

stationary while the second solvent phase passes through the

stationary phase solvent.

Animation

Animation

Rotar

(Instrumentation)

Cost Effective: 10-20%

Efficient: 99.5 – 99.8%

Faster: 2-3 times

Flexible

Easy Scale Up

One single separation /purification techniqueadapted to samples rangingfrommg to several kg.

Bousquet et al*.EPA and DHA from micro algal oil used heptane as the stationary phase and acetonitrile/ water (3% v/v) as the mobile phase.

*O. Bousquet, N. Sellier, and F. L. Goffic, Chrotographia, 39, 40–44 (1994).

Advantages of CPC/CCC & Application

Enzymatic Separations

Long and highly unsaturated

fatty acids show high

resistance for lipases at attack

on ester bonds due to large

stearic hindrance and thus

method is widely used in

separation of 3-PUFA from

marine oils.

Bottino et al.* have illustrated themechanism of resistance of lipasestoward long-chain o3-PUFA inmarine oils.

*N. R. Bottino, G. A. Vandenberg, and R. Reiser, Lipids, 2, 489–493 (1967)

Pure Urea crystallizes intetragonal structure withchannels of 5.67 A° diameter

Crystal channel diameter: 5.6 A0

Fatty acid equivalent diameter: 6.5 – 13.5 A0

Urea complexation

In presence of long

straight-chain molecules,

it crystallizes in hexagonal

structure with inner

channels of 8–12 A°

diameter

Source: A. E. Smith, Acta Cryst., 5, 224–235

TAGs + Alc. KOH

Saponifiable

Unsaponifiable

Almost pure FFAs and Subjected to

Urea Complexation

FFAs

+

Alcohol

Urea

UIC

Two fraction are developed and then UIC fraction is evaporated to separate urea and fatty acids

Step: 1

Step: 2

Urea fractionation has been most frequently used for thepurification of fatty acids from fish oil [1] and from several vegetaloils like, blackcurrant oil [2] borage oil [3]; rapeseed oil [4].

Sources:

1. Ratnayake, WMN et al., Fat Sci Technol, 90, 381, 1988.

2. Traitler H et al., J. Am Oil Chem Soc, 65, 755, 1988.

3. Shimada Y et al., J Am Oil Chem Soc, 75, 1539, 1988.

4. Hayes DG et al., J Am Oil Chem Soc, 75, 1403, 1998.

Advantages & Applications

Crystallization is technically not feasible to separate high unsaturatedfatty acids from moderately high unsaturated fatty acids. e.g.: C18:3 &C18:5 can't easily separated as they have negative melting point, insuch cases UIC is robust tool.

By - products

Component Sunflower Cottonseed Soybean

Unsaponifiables, % 39 42 33

Tocopherols, % 9.3 11.4 11.1

a-tocopherol, % 5.7 6.3 0.9

Sterols, % 18 20 18

Stigmasterol, % 2.9 0.3 4.4

Gums Deodorizer distillate (DOD) Waxes Soap Stock

Deodorizer distillate (DOD)

Tocols from Deodorizer distillate (DOD)

Tocols = Tocopherols + Totrienols

A combination of molecular distillation (MD), ethanol

fractionation, chemical alcoholysis, and ion-exchange

chromatography

MD may not produce a high purity tocopherol because very

similar molecular weights of Tocols and Sterols

By Ethanol fractionation purity can be enhanced as sterols are

insoluble, whereas Tocopherols are soluble in ethanol.

Esterification of alcohols

Series of Distillation

FAME

Tocols

Esters Residues

Barnicki et al. *

Patented a solvent less process, comprised of an esterificationreaction to reduce the volatility of alcohols followed by a series ofdistillation steps, where components boiling higher and lower thantocopherols are separated from tocopherols.

*S. D. Barnicki, C. E. Sumner, Jr., and H. C. Williams, U.S. Patent 5,512,691, 1996.

Sterols are polycyclic alcohols,

Unsaponifiable fraction & quality

depends on temperature, duration,

quantity of stripping steam, and

the extent of vacuum

Phytosterols from DOD

•Either by hydrolysis or trans-esterification

Breaking of ester bonds

•Re-esterification of Phytosterols occurs during methyl ester

Trans-esterification •Phytosterols

content up to 50% by removing FAMEs

Distillation

Instead of Distillation we can go for Crystallization

(physical), solvent extraction (chemical), or crystallization

with additives via adduct formation and separation

(physicochemical)

Organic solvents or solvent mixtures composed of low- and

high-polarity solvents and water are used for crystallization

of Phytosterols

Quality & Quantity of feed and final product are the majorparameters for selection of isolation techniques

Present day demand for a variety of fatty acids, including long-chain polyunsaturated fatty acids (PUFA), may require acombination of different processes and further steps ofseparation

Bulk properties as well as molecular properties both should becombined to isolate the product within the specifications

Good amount of experimental work & processes feasibilityanalysis is required in order implement novel techniques onindustrial scale

CCC/CPC techniques can be used for isolation of oleo-chemicalsat commercial scale

Conclusion

Carbon

No.Chloroform Benzene Cyclohexane Acetone

Ethanol

95%

Acetic

acidMethanol

10 3260 3980 3420 4070 4400 5670 5100

12 830 936 680 605 912 818 1200

14 325 292 215 159 189 102 173

16 151 73 65 53.8 49.3 21.4 37

18 60 24.6 24 15.4 11.3 1.2 1

Solubility

Volatility

Component Mol. Wt. Rel. Volatility

Fatty Acids 280 2.5

Squalene 411 5

Tocopherols 410 1

Sterols 415 0.6

Sterols esters 675 0.04

TAGs 885 small

Systematic Name Trivial NameShorthand

designation

Molecular

wt.

Melting point

(0C)

Dodecanoic Lauric 12:00 200.3 44.2

Tetradecanoic Myristic 14:00 228.4 53.9

Hexadecanoic Palmitic 16:00 256.4 63.1

Heptadecanoic Margaric

(Daturic)

17:00 270.4 61.3

Octadecanoic Stearic 18:00 284.4 69.6

Eicosanoic Arachidic 20:00 312.5 75.3

Docosanoic Behenic 22:00 340.5 79.9

Tetracosanoic Lignoceric 24:00 368.6 84.2

Hexacosanoic Cerotic 26:00 396.7 88

Melting Point (M.Wt.)

Systematic Name Trivial NameShorthand

designation

Molecular

wt.

Melting point

(°C)

Cis-6-

octadecenoicPetroselinic 18:1 (n-12) 282.4 30

Cis-9-

octadecenoicOleic 18:1 (n-9) 282.4 16.2

Tr-9-octadecenoic Elaidic tr18:1 (n-9) 282.4 43.7

Cis-11-

octadecenoic

Vaccenic

(Asclepic)18:1 (n-7) 282.4 39

9,12-

octadecadienoic

Linoleic Acid18:2(n-6) 280.4 -5

6,9,12-

octadecatrienoic

G-linolenic

Acid18:3(n-6) 278.4

Melting Point (Unsaturation)