LIPID ANALYSIS(Ch.13-14)karaali/lipidanalysis.pdf · before analysis. 3- Derived lipids: These are substances derived from lipids and are soluble in lipid-extracting solvents( i.e.

LIPI

D A

NALY

SIS(

Ch.1

3-14

)I.

Qua

ntificat

ion

of li

pid

compo

nent

s

a.Ex

trac

tion

s (we

t vs

dry

;so

lid-l

iqvs

liq-l

iq.)

b.In

dire

ct m

etho

ds(N

MR,

X r

ays,

Con

duct

ance

)

II.

Ass

ays

for

furt

her

inve

stigat

ions

on

extr

acte

d oil

a.Pu

rity

crite

ria:

give

ind

icat

ions

of

wholes

omen

ess

or

adulte

ration

(ph

ysical:n

D20,d

ensity

; ch

emical:

IV,

SV,

FAM

E, s

tero

ls,

toco

pher

ols

compo

sition

s )

b.Q

ualit

y cr

iter

ia:

indica

tes

good

qua

lity

or d

eter

iora

ted

quality

b.1.

MIU

(M

oist

ure,

Impu

rities

, Uns

apon

ifiabl

es)

b.2.

Ranc

idity

: H

ydrt

olyt

ic:A

V an

d ff

a

Oxi

dative

:PV,

TBA,T

OTO

X e

tc

b.3.

Sto

rage

sta

bilit

y(she

lf-l

ife)

Lipi

ds:F

ats

and

fat

like

com

poun

ds(n

otpo

lym

ers)

that

ar

e so

lubl

ein

no

n-po

lar

orga

nic

solv

ents

(pet

role

umet

her)

, but

inso

lubl

e in

H2O

. Lip

ids

have

:

1. Cu

linar

y im

port

ance

: Gre

at e

ffec

ts in

pre

para

tion

of

food

ssi

mnc

e th

ey p

rovi

dehe

at t

rans

fer

med

ia, c

arry

flav

ours

, and

con

trib

ute

to p

alat

abili

ty(e

atin

g qu

alit

y)of

foo

ds.

2. N

utri

tion

al i

mpo

rtan

ce:

Prov

ide

bulk

of

calo

ries

in

diet

(1

g: 9

kca

l).

For

heal

thy

diet

s, %

30 p

erce

nt o

f to

tal c

alor

ies

have

to

com

e fr

om li

pids

.

3.

Phys

iolo

gica

l im

port

ance

: Th

ey

are

vita

l fo

r

bi

olog

ical

fu

ncti

onin

g of

ce

lls,

sinc

e th

ey

cont

ain

esse

ntia

l fat

ty a

cids

and

oil-

solu

ble

vita

min

s.

“LIP

ID”

1.Fa

tty a

cids

2.N

eutra

l fat

s and

oils

3.W

axes

4.Ph

osph

olip

id5.

Ster

ols

6.Fa

t sol

uble

vita

min

s

Lipi

ds in

foo

ds o

ccur

in t

hree

diff

eren

t fo

rms:

1-Si

mple

lipids-

Fats

,oils

are

triglyc

erides

-The

y ar

e th

e es

ter

of

fatt

y ac

ids

with

glyce

rol(a

ll ed

ible

fats

an

d oils); wa

xes

are

este

rs of

fa

tty

acids

with

lon

ger

chain

alco

hols.

2-Co

mpo

site

lipids:

Con

tain

compo

nent

s ot

her

than

lip

ids.

Ex:P

hosp

holip

ids,

Lipo

prot

eins

,Glyco

lipids:

Th

ese

are

foun

d in

boun

d fo

rm

with

ot

her

cons

titu

ents

;th

eir

ana

lyse

s is h

arde

rth

an s

imple

lipids,

bec

ause

you

hav

e to

hyd

rolyse

the

molec

ule

befo

re a

nalysis.

3-Der

ived

lipids:

The

se a

re s

ubst

ance

s de

rive

d fr

om

lipids

and

are

solubl

e in l

ipid-e

xtra

cting

solven

ts(

i.e.

fat

solubl

e pigm

ents

, st

erols,

ant

ioxida

nts

like

toco

pher

ols

and

Vita

mins

A,D

, E,

K(f

at-s

olub

le

vita

mins).T

hey

are

not

true

lipids

but

are

foun

d in

lipid

phas

e.

They

be

have

lik

e lip

ids.

In

mos

t an

alys

es y

ou w

ill g

et t

hem in

the

lipid f

ract

ion.

Fatty

Aci

ds

OR

C

O

H

#1 C

arbo

nA

cid

Gro

up

OR

C

O

H

Non

-pol

ar E

nd -

Hyd

roph

obic

End

(

Fat-s

olub

le ta

il) Po

lar E

nd -

Hyd

roph

ilic

End

Satu

rate

d Fa

tty A

cids

CH

3

CH

2

CH

2

CH

2

CH

2

CH

2

CH

2

C

OH

O 12

45

67

83

Oct

anoi

cA

cid

Uns

atur

ated

Fat

ty A

cids

CH

3

CH

2

CH

2

CH

2

CH

2

CH

2

CH

2

C

OH

O 12

45

67

83

CH

3

CH

2

CH

2

CH

2

CH

2

CH

2

CH

2

C

OH

O 12

45

67

83

3 -O

cten

oic

Aci

d

3, 6

-O

ctad

ieno

icA

cid

Shor

t han

d:8:

1 (∆

3)

8:2

(∆3,

6)

Cis

9 -O

ctad

ecen

oic

Aci

d (o

leic

)

Tran

s 9 -

Oct

adec

enoi

cA

cid

(ela

idic

acid

)

OC

H3(

CH

2)7

C

C

(C

H2)

7

C

OH

HH

910

OC

H3(

CH

2)7

C

C

(C

H2)

7

C

OH

H

H

Cis

And

Tra

ns F

atty

Aci

ds

Poly

unsa

tura

ted

Fatty

Aci

ds

Lino

leic

acid

:C

is, c

is, 9

, 12

-Oct

adec

adie

noic

acid

Lino

leni

cac

id:

Cis

, cis

, cis

9, 1

2, 1

5 -O

ctad

ecat

rieno

icac

id

Ara

chid

onic

acid

:C

is, c

is, c

is, c

is5,

8, 1

1, 1

4 -E

icos

atet

raen

oic

acid

Lino

leic

Aci

d

Lino

leni

cA

cid

Ara

chid

onic

Aci

d

Nat

ural

ly-o

ccur

ring

fatty

aci

ds

R

CH

2

CH

C

H

CH

2

CH

C

H

CH

2

C

OH

O

76

54

3

1.C

isfo

rm

2.N

ot c

onju

gate

d --

-iso

late

d do

uble

bon

d.

3.Ev

en n

umbe

red

fatty

aci

ds.

FAT

AN

D O

ILS

Mos

tly T

rigly

cerid

es:

O

H2C

O

H

HC

O

H

H2C

O

H

HO

C

R

HO

C

R

HO

C

R

OO O

H2C

O

C

R

O OH

2C

O

C

R

HC

O

C

R

++

3 H

2O

Gly

cero

l 3

Fatty

Aci

ds

GLY

CER

IDES

Mon

ogly

cerid

e(α

-mon

oste

arin

)D

igly

cerid

e(α

, α' -

dist

earin

)

H2C

O

H

HC

O

H

H2C

O

O C

(CH

2)16

CH

3

H2C

O

HC

O

H

H2C

O

O C

(CH

2)16

CH

3

C

(CH

2)16

CH

3

O

Trig

lyce

ride

(β-p

alm

ityld

iste

arin

)

H2C

O

HC

O

H2C

O

O C

(CH

2)16

CH

3

C

(CH

2)16

CH

3

O O C

(CH

2)14

CH

3

(C18

)

(C16

)

(C18

)

α-o

leod

ipal

miti

n

1 -o

leod

ipal

miti

n

Ole

ic

Palm

itic

Palm

itic

OPP

α-L

inol

eyld

iole

in

1 -L

inol

eyld

iole

in

Ole

ic

Lino

leic

Ole

icLO

O

FATS

AN

D O

ILS

AR

E PR

IMA

RIL

Y T

RIG

LYC

ERID

ES (9

7-99

%)

Veg

etab

le o

il -

68%

of w

orld

supp

lyC

ocoa

but

ter -

solid

fat

Oil

seed

s -liq

uid

oil

Ani

mal

fat -

28%

(fro

m H

ogs a

nd C

attle

)

Mar

ine

oil -

4%W

hale

oil

cod

liver

oil

Fatty

Aci

dC

ompo

sitio

ns (%

) of F

ats a

nd O

ils

43

63

82

610

36

123

4414

1018

116

2611

412

16:1

71

18:0

156

32

18:1

297

1824

18:2

22

5354

Fatty

Aci

dsB

utte

rC

ocon

utC

otto

nsee

dSo

ybea

n

18:3

28

WA

XE

SFa

tty a

cids

+ L

ong

chai

n al

coho

l

Impo

rtant

in fr

uits

:

1.N

atur

al p

rote

ctiv

e la

yer i

n fr

uits

, veg

etab

les,

etc.

2.A

dded

in so

me

case

s for

app

eara

nce

and

prot

ectio

n.

Bee

swax

(myr

icyl

palm

itate

), Sp

erm

acet

i (ce

tylp

alm

itate

)

OC 3

0H61

O

C

C

15H

31

OC

16H

33

O

C

C15

H31

PHO

SPH

OLI

PID

Leci

thin

(pho

spha

tidyl

chol

ine)

O O

H2C

O

C

R

O

R

C

O

CH

H2C

O

P

O

C

H2

C

H2

N

+ CH

3

CH

3

CH

3

O_

Phos

phat

idic

Aci

dC

holin

e

STER

OLS

Mal

e &

fem

ale

sex

horm

ones

Bile

aci

dsV

itam

in D

Adr

enal

cor

ticos

tero

ids

Cho

lest

erol

Ph

ytos

tero

ls(i.

e. Β

-sito

ster

ol)

HO

H3C

H3C

H3C

CH

3

CH

3

12 3

45

678

910

1112

1314

151617

182122

19

20

FAT

SOLU

BLE

VIT

AM

INS

Vita

min

A:

CH

2O

HC

H3

CH

3

CH

3

CH

3H

3C

12

34

5

67

89

Vita

min

D2:

Vita

min

E:

HO

CH

2

HH

H3

CH

3C

CH

3

CH

3 CH

3

OR

1

R2

HO

R3

CH

3 (CH

2CH

2CH

2CH

2)2C

H2C

H2C

H2C

H(C

H3)

2

CH

3

Fat

cont

ents

of s

elec

ted

food

s:

Liqu

id v

eget

able

oils

≅10

0%

Mar

gari

ne a

nd b

utte

r ≅

80%

→th

ey a

re e

mul

sion

s of

lipi

d in

wat

er

Alm

onds

:%55

, W

alnu

ts:%

65, C

erea

ls:3

-5%

Sunf

lowe

r se

eds,

hul

led

: ~6

0%

Choc

olat

e:~3

5% ,

Milk

:37

%, M

ilk p

owde

r : 2

7.5%

Tuna

fis

h: 4

% (

but

ther

e ar

e so

me

fatt

y fi

sh li

ke

anch

ovie

s co

ntai

ning

15%

fat

)

Ia. G

ener

al p

roce

dure

for

sol

vent

ext

ract

ions

:

1.Sam

ple

Prep

arat

ion:

Plant

or

anim

al t

issu

es s

hould

be h

omog

enized

by

grind

ing,

blen

ding

etc

, sinc

e pa

rticle s

ize

is

impo

rtan

t; it

has

to

be

idea

lpa

rticle

size

for

max

imum

ex

trac

tion

yields

(i.e.

0.4m

mφ)

. th

en it ha

s to

be

dr

ied

befo

re ex

trac

tion

, be

caus

e if a

ny m

oist

ure

is let

sta

y in t

he s

amples

, it int

erfe

res

with

ex

trac

tion

, an

d th

e so

lven

t's

extr

action

ca

pabi

lity

decr

ease

s.Also

hydr

olys

is is ne

eded

to

br

eak

down

an

y bo

und

lipids,

eithe

r by

aci

ds(H

Clor

H2S

O4)

or b

y en

zym

es l

ike

Taka

dias

tase

.

2.

Solve

nt

select

ion:

Yo

u ne

ed

diff

eren

t ty

pes

of

solven

ts

for

extr

acting

diffe

rent

typ

es o

f lip

ids:

(Chlor

ofor

m+m

etha

nol)

is id

eal

for

lipop

rote

ins,

wh

erea

s pe

troleu

met

her

frac

tion

s(C6

an

d C5

) ar

e idea

l fo

r tr

ue

lipid

s (triyg

lyce

ride

s),

and

ethy

l et

her

for

extr

acting

ox

idized

lip

ids.

So

met

imes

a t

erna

ry m

ixtu

re w

orks

bet

ter

(CHCl

3: m

etha

nol:H2O

) fo

r ex

trac

tion

fro

m a

nimal

tiss

ues;

wat

er-s

atur

ated

but

anol f

or

cere

al lipid

s .

LIPI

D C

ON

TEN

T A

NA

LYSE

S

1.G

ravi

met

ric M

etho

ds

(1) W

et e

xtra

ctio

n -R

oese

Got

tlieg

b&

Moj

onni

er.

(2) D

ry e

xtra

ctio

n -S

oxhl

etM

etho

d.

2.V

olum

etric

Met

hods

(Bab

cock

, Ger

ber M

etho

ds)

1. G

ravi

met

ric M

etho

ds

(1)

Wet

Ext

ract

ion

-Roe

seG

ottli

eb &

Moj

onni

er.

For M

ilk:

1) 1

0 g

milk

+ 1

.25

ml N

H4O

H m

ix. s

olub

ilize

spr

otei

n an

d ne

utra

lizes

.

2) +

10

ml E

tOH

-sha

ke.

Beg

ins e

xtra

ctio

n, p

reve

nts

gela

tion

of p

rote

ins.

3) +

25

ml E

t2O

-sh

ake

and

mix

.

4) +

25

ml p

etro

leum

eth

er, m

ix a

nd sh

ake.

(2)

Dry

Ext

ract

ion

-Sox

hlet

Met

hod.

Sam

ple

in th

imbl

e is

con

tinuo

usly

ext

ract

ed w

ith e

ther

us

ing

Soxh

letc

onde

nser

. A

fter e

xtra

ctio

n, d

irect

m

easu

rem

ent o

f fat

-eva

pora

te e

ther

and

wei

gh th

e fla

sk.

Indi

rect

mea

sure

men

t -dr

y th

imbl

e an

d w

eigh

thim

ble

and

sam

ple.

2. V

olum

etric

Met

hods

(Bab

cock

, Ger

ber M

etho

ds)

Theo

ry:

1.Tr

eat s

ampl

e w

ith H

2SO

4 or

det

erge

nt.

2.C

entri

fuge

to se

para

te fa

t lay

er.

3.M

easu

re th

e fa

t con

tent

usi

ng sp

ecia

lly c

alib

rate

d bo

ttles

.

Met

hods

:1.

Kno

wn

wei

ght s

ampl

e.2.

H2S

O4

-dig

est p

rote

in, l

ique

fy fa

t.3.

Add

H2O

so th

at fa

t will

be

in g

radu

ated

par

t of b

ottle

.4.

cent

rifug

e to

sepa

rate

fat f

rom

oth

er m

ater

ials

com

plet

ely.

Typ

es o

f Ex

trac

tion

:a1

.. "D

ry E

xtra

ctions

":Th

ese

are

basi

cally

sol

id-

liqui

dty

pe o

f ex

trac

tion

s,an

d th

e fi

nal

resu

lts

are

grav

imet

ric.

Th

ey

may

be

ei

ther

ba

tch,

m

ulti

ple-

batc

h,

cont

inuo

us

or

coun

terc

urre

nt

extr

acti

ons.

Th

ere

are

also

au

tom

ated

pr

oces

ses,

like

FOSS

LET

extr

acto

rs, w

here

the

pr

edet

erm

ined

num

ber

of s

tage

s is

(4

-5),

and

cart

ridg

esar

e us

ed t

o co

ntai

n th

e sa

mpl

e.In

co

mpl

ex m

atri

ces,

dig

esti

on m

ay b

e re

quir

ed t

o lib

erat

e bo

und

lipid

s by

hy

drol

ysis

be

fore

star

ting

the

ext

ract

ion.

Solven

t Ex

trac

tion

Met

hods

:a.

1.1.

Cont

inuo

us(G

oldf

ish)

or

semicon

tinu

ous(Sox

hlet

) or

disc

ontinu

ous(M

ajon

nier

) so

lven

t ex

trac

tion

met

hods

: Solve

nt

from

a

boiling

flas

k co

ntinuo

usly f

lows

ove

r th

e sa

mple

held in

a th

imbl

e.Fa

t co

nten

t is m

easu

red

by w

eigh

t of

fa

t re

mov

ed.

a.1.

2.El

evat

ed

pres

sure

/Tem

pera

ture

met

hods

: SFE

(Sup

ercr

itical

fluid

extr

action

s):inc

e it

avoids

th

e us

e of

ha

zard

ous

orga

nic

solven

ts,

and

uses

ine

rt,

inex

pens

ive,

and

non

toxi

c ga

ses

like

CO2, i

t is g

aining

mor

e po

pularity

.

a.2.

“W

etno

nsolve

nt e

xtra

ctions

":

are

pref

erre

d f

or

liqui

d sa

mpl

es, s

ince

the

y in

volv

e liq

-liq

extr

acti

on

(par

titi

onin

g of

mat

eria

l bet

ween

two

imm

isci

ble

liqui

ds)

and

yiel

d vo

lum

etri

c re

sult

s.

Ex: G

erbe

ran

d Ba

bcoc

k ty

pe o

f ex

trac

tion

of

milk

are

ex

ampl

esof

wet

ext

ract

ion.

In

milk

, the

fat

glo

bule

s ar

e pr

esen

t in

the

for

m o

f an

em

ulsi

onof

“oil

in w

ater

”, su

rrou

nded

by

a th

in f

ilm o

f pr

otei

n wh

ich

has

to b

e re

mov

ed o

r br

oken

by

acid

tre

atm

ent

befo

re s

epar

atin

g fa

t fr

om w

ater

pha

se.

Gerb

erPr

oced

ure:

11

ml m

ilk a

nd 1

ml i

soam

ylal

coho

l are

m

ixed

to

impr

ove

phas

e se

para

tion

, by

incr

easi

ng t

he w

ater

ph

ase.

Then

H2S

O4

(Sp.

gr.

:1.8

2 fo

r br

eaki

ng d

own

the

lipid

-pro

tein

bar

rier

) is

adde

d an

d m

ixtu

re is

cen

trif

uged

fo

r 5

min

,sub

mer

ged

in H

2O b

ath

(65°

C, 5

min

). V

olum

es

of t

wo r

espe

ctiv

e ph

ases

giv

es f

at-w

ater

rat

io, o

r th

e up

per

laye

r gi

ves

fat

cont

ent.

I.b.

Ind

irec

t M

etho

dsfo

r Q

uant

ific

atio

n of

Lip

ids

Espe

cial

ly g

enet

icis

ts i

n pl

ant

bre

edin

g re

sear

ch s

tudi

es

requ

ire

“non

-des

truc

tive

”ty

pes

of li

pid

anal

yses

. Fo

r th

is

purp

ose,

man

y “in

stru

men

tal s

yste

ms”

are

deve

lope

d:

*NM

R:Lo

w re

sool

utio

nnu

clea

rm

agne

tic

reso

nanc

e:Th

e m

easu

red

NM

R va

lue

is r

elat

ed t

o th

e to

tal

num

ber

of

prot

ons

in o

il fr

acti

on o

f se

ed ,

inde

pend

ent

of p

roto

ns in

no

n-oi

l fra

ctio

n, s

ince

pro

tons

ha

ve a

dif

fere

nt m

agne

tic

reso

nanc

e in

oil

and

in

wate

r ph

ase.

It

give

s ve

ry g

ood

appr

oxim

atio

n of

lipi

d le

vels

. Cal

ibra

tion

tab

les

or c

urve

s ar

e us

ed f

or d

iffe

rent

H 2

0 le

vels

(fig

. 13-

8).

*X-r

ay

abso

rbti

on:

Is

an

appr

oved

m

etho

d fo

r m

eat

carc

ass

sam

ples

. Le

an (

yağs

ız)

mea

t ab

sorb

s m

ore

x-ra

y th

an f

atty

tis

sues

(car

cass

-lipi

ds)

Cond

ucta

nce:

Lean

mea

t is

20-

fold

bet

ter

cond

ucto

r fo

r el

ectr

ical

cur

rent

than

fat

.

Den

sito

met

ric

Met

hods

:D

ensi

ty

of

seed

s (i.

e.

Flax

seed

) is

pro

port

iona

l to

its

oil c

onte

nt. L

ikew

ise,

de

nsit

y of

ext

ract

ing

solv

ent

chan

ges

prop

orti

onal

to

the

fat

cont

ent

it

has

extr

acte

d(i.e

.per

chlo

roet

hyle

ne

in

FOSS

LET)

. Fo

r m

eat

, the

re is

a m

etho

d us

ing

a cy

lindr

ical

con

tain

er

with

a p

isto

n pl

ate

that

hol

ds m

eat

unde

r co

nsta

nt

hydr

aulic

pre

ssur

e. M

easu

red

impr

essi

on d

epth

is

rela

ted

to f

at c

onte

nt.

Refr

acto

met

ry:

η D o

f th

e m

ixtu

re (

brom

onap

htal

ene

and

fat

extr

acte

d by

it f

rom

cho

cola

te)

is u

sed

for

appr

oxim

atio

n of

fat

in c

hoco

late

.

II. A

ssay

s on

Ext

ract

ed L

ipid

s-F

AT

CHA

RACT

ERIS

ATI

ON

(Ch.

14):

II.a

. Pu

rity

Crite

ria:

For

auth

enti

city

tes

ting

an

d de

term

inat

ion

of

adul

tera

tion

. M

ost

anal

ytic

al v

alue

s of

cer

tain

oils

and

fat

s ar

e qu

ite

char

acte

rist

ic:

a.1.

Phys

ical

purity

crite

ria

are

refr

acti

ve

inde

x,

mel

ting

po

int,

co

lour

,de

nsit

y,.

By

mea

suri

ng t

hese

cri

teri

a, w

e wi

ll fi

nd v

alue

s wi

thin

cer

tain

acc

epte

d lim

its

and

rang

es.

But

thes

e al

one

are

not

eno

ugh

for

auth

enti

city

ch

arac

teri

sati

on.

Col

or:E

ither

Lov

ibon

d co

lor s

cale

usi

ng

stan

dard

red-

yello

w-b

lue

filte

rs, o

r de

term

inat

ion

of “

phot

omet

ric c

olor

inde

x-PC

I)*

is re

quire

d fo

r obj

ectiv

e de

scrip

tion

of

colo

r.

Ref

ract

ive

Inde

x:R

I may

als

o be

use

d fo

r co

ntro

lling

hyd

roge

natio

n, si

nce

RI w

ill

decr

ease

line

arly

as u

nsat

urat

ion

decr

ease

s.

Mel

ting

poin

t, sm

oke,

flas

h an

d fir

e po

ints

)(4.

56)

(2.41

)(7.

69)

(29.1

*67

062

055

046

0A

AA

APCI

−+

+=

Effe

cts o

f Dou

ble

Bon

ds o

n th

e M

eltin

g Po

ints

16:0

6016

:11

18:0

6318

:116

18:2

-518

:3-1

120

:075

F. A

.M

. P. (

0 C)

20:4

-50

M.P

.

# D

oubl

e bo

nds

a.2.

Ch

emical

purity

cr

iter

ia:

Thes

e ar

e ve

ry

char

acte

rist

icfo

r ea

ch o

il an

d fa

t ty

pes:

•Sa

poni

fica

tion

valu

em

easu

ring

ave

rage

f.a

. m

olec

ular

we

ight

and

cha

in le

ngth

•I

odin

e va

lue

mea

suri

ngle

vel o

f un

satu

rati

on

•Fa

tty

acid

com

posi

tion

(FA

ME)

,

•In

divi

dual

ste

rola

nd t

ocop

hero

l com

posi

tion

s.

Wit

h an

inte

grat

ed e

valu

atio

n of

res

ults

of

puri

ty

crit

eria

ana

lyse

s,we

can

be

sure

of

iden

tity

and

sour

ce o

f fa

ts a

nd o

ils(R

efer

ence

met

hods

in

AO

CS,I

UPA

C an

d D

GF).

1. S

apon

ifica

tion

Val

ue(1

4.3.

9)

Sapo

nific

atio

n-h

ydro

lysi

s of e

ster

link

ages

und

er

alka

line

(KO

H)c

ondi

tion.

O

C

ROO C

R

C

RO

H2C

O

HC

O

H2C

O

KO

H

HHH

H2C

O

HC

O

H2C

O

R

C

OK

+3

+3

Milk

Fat

210-

233

Coc

onut

Oil

250-

264

Cot

ton

Seed

Oil

189-

198

Soyb

ean

Oil

189-

195

Fat

Sapo

nific

atio

nV

alue

Lard

190-

202

Sapo

nific

atio

nV

alue

of F

ats a

nd O

ils

Sapo

nific

atio

nV

alue

=-m

gs o

f KO

H r

equi

red

to sa

poni

fy1

g of

fat.

1.5

g in

250

ml E

rlen

mey

er.

2.50

ml K

OH

in E

rlen

mey

er.

3.B

oil f

or sa

poni

ficat

ion.

4.T

itrat

e w

ith H

Clu

sing

phe

nolp

htha

lein

.5.

Con

duct

bla

nk d

eter

min

atio

n.

B -

ml o

f HC

lreq

uire

d by

Bla

nk.

S -m

l of H

Clr

equi

red

by S

ampl

e.

SP#

= 56

.1(B

-S) x

N o

f HC

l

Gra

m o

f Sam

ple

Sapo

nific

atio

nV

alue

Det

erm

inat

ion

SV is

an

inde

x of

the

mea

n m

olec

ular

wei

ght

of t

he

tria

cylg

lyce

rols

in s

ampl

e oi

l or

fat.

The

mea

n m

olec

ular

wei

ght

of t

riac

ylgl

ycer

ols

when

di

vide

d by

thr

ee w

ill g

ive

an a

ppro

xim

ate

mea

n m

olec

ular

wei

ght

of f

atty

aci

ds p

rese

nt.

2. I

odin

e N

umbe

r (14

.3.8

)

Iodi

ne v

alue

(IV)

: Hal

ogen

s ad

d to

dou

ble

bond

s.

IV is

the

num

ber

of g

ram

s of

iodi

ne t

hat

adds

to

100

gram

of

oil.

If 1

g o

f the

fat a

dsor

bs1.

5 g

of io

dine

, the

n IV

= 15

0.

It is

a m

easu

re o

f sa

tura

tion

-uns

atur

atio

n.

Iodi

ne V

alue

= (m

l of N

a 2S 2

O3

volu

me

for

blan

k -

ml o

f N

a 2S 2

O3

volu

me

for

sam

ple)

×N

of N

a 2S 2

O3

×0.

127g

/meq

×10

0

Wei

ght o

f Sam

ple

(g)

CH

C

HC

H

CH

Cl

IIC

lIo

dine

chl

orid

e+

ICl

KI

KC

l

I 2

I 2

Na

2S

2O

3N

a2

S4

O6

NaI

+

+2

2+

+

Exc

ess u

nrea

cted

ICl

Iodi

ne V

alue

Det

erm

inat

ion

Iodi

ne N

umbe

rs o

f Sel

ecte

d F.

A.

Palm

itole

icA

cid

195

Ole

ic A

cid

186

Lino

leic

Aci

d2

173

Lino

leni

cA

cid

326

1

Fatt

y A

cids

# of

Dou

ble-

bond

sIo

dine

#

Ara

chid

onic

Aci

d4

320

If m

olec

ular

wei

ght a

nd io

dine

num

ber o

f f.a

. isk

now

n,

we

can

cal

cula

te th

e nu

mbe

r of d

oubl

e bo

nds:

IV/M

W

Dur

ing

oxid

atio

n,

ther

e wi

ll be

a

decl

ine

in

unsa

tuar

tion

and

the

refo

re in

IV:

The

rang

es o

f Io

dine

Val

uefo

r ol

ive

oil,

whic

h is

an

inde

x of

its

degr

ee o

f un

satu

rati

on,

is 7

5-94

in t

he C

odex

. F

or I

taly

thi

s va

lue

fluc

tuat

es

betw

een

79 a

nd 8

8, b

ut f

or T

unis

iath

e ra

nge

is

80-9

7.Th

ere

may

be

such

reg

iona

l or

clim

atic

va

riat

ions

, th

eref

ore,

ev

en

thou

gh r

esul

ts a

re

quit

e m

eani

ngfu

l, so

met

imes

add

itio

nal

test

s ar

e re

quir

ed f

or f

inal

con

firm

atio

n.

GC

Ana

lysi

s for

Fat

ty A

cids

(14.

6.1)

1.Ex

tract

fat.

2.Sa

poni

fy(h

ydro

lysi

s und

er b

asic

con

ditio

n).

3.Pr

epar

e “f

atty

aci

d m

ethy

l est

ers”

or F

AM

E (b

y So

dium

met

hoxi

de-

CH

3ON

a or

BF3

-met

hano

l rea

gent

).

4.C

hrom

atog

raph

y of

met

hyl e

ster

s.

5.D

eter

min

e pe

ak a

reas

of f

atty

aci

ds.

(Fat

ty a

cids

are

iden

tifie

d by

thei

r spe

cific

rete

ntio

n tim

es).

6.C

ompa

re w

ith re

spon

se c

urve

of s

tand

ard .

14

18:1

18:2

2018

:3

22

21:1

2416

18

Tim

e

Res

pons

e

For

anal

ysin

gFA

ME:

Th

e FA

ME

are

carr

ied

in a

nd o

ut o

f th

e GC

Col

umn

by

the

carr

ier

gas

and

each

will

leav

e th

e co

lum

n on

ly a

fter

a

spec

ific

ret

enti

on t

ime,

Rt, w

hich

is d

epen

dant

on

the

chai

n le

ngth

and

deg

ree

of s

atur

atio

n of

the

fat

ty a

cid.

Ge

nera

lly, a

s ca

rbon

num

ber

as w

ell a

s do

uble

bon

ds in

f.

atty

aci

ds i

ncre

ase,

the

rete

ntio

n ti

me

Rt w

ill

also

in

crea

se.

FAM

E:Th

e fa

tty

acid

com

posi

tion

det

erm

ined

by

GC

(gas

chr

omot

ogra

phy)

is a

real

"Fi

nger

prin

t" .

It

also

se

rves

fo

r “n

utri

tion

al

labe

lling

”: sa

tura

teds

,mon

ouns

atur

ates

, pol

yuns

atur

ates

, tra

ns f

.a.

etc.

For

auth

enti

city

tes

ting

of

butt

ers,

thi

s an

alys

isby

its

elf

may

not

som

etim

es b

esu

ffic

ient

, si

nce

som

e m

arga

rine

s m

ight

hav

e si

mila

r co

mpo

siti

on o

f FA

ME

with

but

ters

.

In t

his

case

,the

res

pect

ive

ster

ol c

ompo

siti

onsh

ould

als

o be

dete

rmin

ed. S

tero

l ana

lysi

s is

don

e by

TLC

fol

lowe

d by

GC

of

the

unsa

poni

fiab

lefr

acti

on

of l

ipid

s. (

Ther

e ar

e “P

hyto

ster

ols”

-st

igm

aste

rol,

cam

pest

erol

, β-

sito

ster

ol-

inth

e ve

geta

ble

ster

olfr

acti

on, b

ut o

nly

“cho

lest

erol

”in

an

imal

fat

s).

CH

OLE

STER

OL

DET

ERM

INA

TIO

N(1

4.6.

4)C

hole

ster

ol b

y G

LC:s

apon

ify e

xtra

cted

lipi

d,de

rivat

ize

G

C

µg/

ml C

hole

ster

ol

Abs

orpt

ion

at

440

nm

Spec

trom

ertic

Abs

orpt

ion

Stan

dard

Cur

ve o

f Cho

lest

erol

Enzy

mat

ic D

eter

min

atio

n: C

hole

ster

ol O

xida

se H

OO

H2O

2

Cho

lest

erol

Oxi

dase

etc.

+

H2O

2

CH

3OO

CH 3

H2N

NH 2

HN

NH

OC

H 3C

H3O

H2O

Pero

xida

se+

+

0-D

iani

sidi

neO

xidi

zed

0-D

iani

sidi

ne(C

olor

less

)(B

row

n co

lor)

At 4

40 n

m

TRIG

LYC

ERID

E A

NA

LYSI

S

BY

LIQ

UID

CH

RO

MA

TOG

RA

PHY

Soyb

ean

Oil

Solv

ent C

H3C

N/H

FC

olum

n 84

346

(Wat

ers A

ssoc

iate

s)

RES

PON

SE

RET

ENTI

ON

TIM

E

Ole

ate-

cont

aini

ng tr

igly

ceri

des i

n ol

ive

oil

OL

254

:544

O2L

54:4

46

OPL

52:3

46

O3

54:3

48

OSL

54:3

48

O2P

52:2

48

O2S

54:2

50

OPS

52:1

50

Fatt

y A

cid

Com

posi

tion

Tot

al A

cylC

arbo

ns:

Uns

atur

atio

nE

quiv

alen

t Car

bon

Num

ber

OS2

54:1

52

Furt

her

Spec

ific I

dent

ificat

ion

Test

s fo

r So

meF

ats

and

Oils

:( F

or d

etec

tion

of

adul

tera

tion

)

For

butt

er: R

eich

ert-

Mei

ssel

inde

x fo

r bu

tter

is a

mea

sure

of

its

con

tent

of

vola

tile

and

wat

er-s

olub

le b

utyr

ic a

cid;

>2

4.Fo

rol

ive

oil:

Squa

lene

(700

0 m

g/kg

) is

a

very

spe

cifi

c co

nsti

tuen

t of

un

sapo

nifi

able

par

t of

oliv

e oi

l, wh

erea

s in

ot

hers

, it

vari

es b

etwe

en 3

0-40

0ppm

.Fo

r co

tton

seed

oil:

Hal

phen

test

for

dete

ctin

g pr

esen

ce o

f cy

clop

rope

noic

aci

ds w

hich

for

m a

red

col

or o

n he

atin

g wi

th a

myl

alc

ohol

and

CS2

.Fo

r se

sam

esee

d oi

l: Vi

llave

chia

tes

t: u

nsap

onif

iabl

es o

f se

sam

esee

d oi

l onl

y wi

ll fo

rm p

ink

colo

r wi

th f

urfu

ral.

For

coco

butt

er: T

rigl

ycer

ides

C50

and

C54

are

dom

inan

t in

co

cobu

tter

but

not

in C

BE a

nd C

BS.

II b

. Qua

lity

Crit

eria

IIb.

1. M

I U

(M

oist

ure+

Inso

lubl

e Im

puri

ties

)(d

irt,

oil

seed

pa

rtic

les

in

crud

e oi

ls

and

soap

in

re

fine

d oi

ls)+

(uns

apon

ifia

bles

) :

They

are

nonf

atty

cons

titu

ents

of

oils

whic

h in

crea

se

refi

ning

co

sts

for

crud

e oi

ls

.

Ther

efor

e,

ever

y pe

rcen

t of

in

crea

se

in

MIU

co

nten

t br

ings

par

alle

l pr

ice

decr

ease

sin

cru

de o

il pu

rcha

ses.

Als

o so

ap>5

ppm

lef

t in

ref

ined

oils

will

be

det

rim

enta

l to

oil

qual

ity

.II

b.2

Fat

ranc

idit

yan

d st

abili

ty

Ranc

idit

y:“D

evel

opm

ent

of

obje

ctio

nabl

e fl

avou

rsan

d od

ours

resu

ltin

g fr

om l

ipol

ysis

or

lipid

bre

ak-d

own

(eit

her

due

to h

ydro

lyti

c or

to

oxid

ativ

e re

acti

ons)

lead

ing

to lo

wer

cons

umer

acc

epta

nce”

.St

abili

ty:

“Cap

abili

ty t

o re

mai

n fr

esh

or m

aint

ain

fres

h ta

ste+

odou

r”or

“re

sist

ance

to

spoi

lage

dur

ing

stor

age

or

“res

ista

nce

of li

pids

to

ranc

idit

y”

RANCI

DIT

Y(Hyd

rolytic

and

Oxi

dative

)

A-H

ydro

lyti

c Ra

ncid

ity:

Invo

lves

che

mic

al o

r en

zym

atic

(lipa

se

from

mic

roor

gani

sms

or p

lant

cel

ls)

hydr

olys

is o

f fa

ts in

to f

ree

fatt

y ac

ids

and

mon

o or

di-g

lyce

ride

s. R

elev

ant

Test

s:A

V an

d ff

aa.

A.V

.=Th

e nu

mbe

r of

mili

gram

sof

KO

H r

equi

red

to n

eutr

aliz

e 1

gram

of

fat

or o

il.T

and

aw

b.FF

A:

Free

fa

tty

acid

s=

here

, ac

idit

y is

ex

pres

sed

as

perc

enta

geof

wei

ght

of a

spe

cifi

c fr

ee f

atty

aci

d in

a f

at o

r oi

l,thu

s ta

king

into

con

side

rati

on t

he M

.W. o

f th

e in

divi

dual

f.a

.:*

In o

live

oil,

expr

esse

d as

ole

icac

id,

MW

= 1

82*I

n pa

lm o

il, e

xpre

ssed

as

palm

itic

aci

d,M

W =

256

* %

FFA

ole

ic a

cid=

0.5

03 x

Aci

d va

lue

* A

cid

valu

e= 1

.99

x %

FFA

ole

ic a

cid

100

)10

00(

%x

Weight

acids

fatty

MW

xV

FFA

=

AV

=

ml o

f KO

H x

N x

56

Wei

ght o

f Sam

ple

= m

g of

KO

H

Sign

ifican

ce o

f AV

and

ffa:

This

val

ue w

ill e

stim

ate

the

amou

nt o

f cr

ude

oil t

hat

will

be lo

st d

urin

g re

fini

ng, s

o wi

ll m

ean

high

er o

pera

ting

co

sts

for

refi

ning

pla

nts,

and

lead

s to

pri

ce d

ecre

ases

in

crud

e oi

l pur

chas

es.

In r

efin

ed o

ils,

high

val

ues

will

mea

n ei

ther

poo

r re

fini

ng o

r fa

t br

eakd

own

in s

tora

ge o

r us

e. I

n ca

ses

wher

e th

e li

bera

ted

ffa

are

vola

tile

, ff

a wi

ll be

a

mea

sure

of

hydr

olyt

ic r

anci

dity

.

For

Vir

gin

oliv

e oi

ls <

4 %

ole

ic a

cid,

and

for

ref

ined

oliv

e oi

ls <

0.3;

but

for

sho

rter

cha

in f

.a. l

ike

buty

ric,

capr

ic a

nd e

ven

laur

ic (C

4-C1

2), h

avin

g m

uch

lowe

r fl

avou

r th

resh

olds

(as

in c

hoco

late

), th

e pr

oduc

t

will

be in

edib

le a

fter

its

ffa

cont

ent

reac

hes

> 0.0

2 %

.

B-O

xida

tive

Ran

cidi

ty

“LIP

ID O

XID

ATI

ON

“M

ajor

flav

or p

robl

ems i

n fo

od d

urin

g st

orag

e ar

e m

ainl

y du

e to

the

oxid

atio

n of

lipi

d.

Lipi

d O

xida

tion

-fre

e ra

dica

l rea

ctio

ns.

1.In

itiat

ion.

2.Pr

opag

atio

n.

3.Te

rmin

atio

n.

Met

hods

for

Mea

suring

Pre

sent

Sta

tus

of

Lipid

Oxi

dation

:-P

erox

ide

num

ber:

The

amou

nt o

f io

dine

lib

erat

ed(N

a 2S 2

O3

titr

atio

n)fr

om s

atur

ated

KI

solu

tion

by

fat

, exp

ress

sed

in

meq

of

pero

xide

O2/

kg o

f fa

t(L

ea v

alue

=exp

ress

ed in

mill

imol

es o

f O

2).

<10

for

edib

le o

ils; m

easu

res

only

tra

nsie

nt p

rim

ary

oxid

atio

n pr

oduc

ts li

ke p

erox

ides

and

hyd

rope

roxi

des.

-Thi

obar

bitu

ric

acid

val

ue (

TBA

):Th

e pi

nk c

olor

for

med

by

seco

ndar

y ox

idat

ion

prod

ucts

like

mal

onal

dehy

des

with

TBA

re

agen

t i

s m

easu

red

at 5

23 n

m.C

orre

late

s be

tter

wit

h se

nsor

y an

alys

es.

-Ani

sidi

neva

lue:

Ano

ther

co

lor

reac

tion

of

alde

hyde

s(al

kena

ls

and

dien

als-

seco

ndar

y pr

oduc

ts

of

oxid

atio

n) w

ith

P-an

isid

ine.

-Tot

oxva

lue

:2

tim

es p

erox

ide

valu

e +

anis

idin

eva

lue

= To

tal

oxid

atio

n pr

oduc

ts (

2 is

bec

ause

per

oxid

es h

ave

2 ti

mes

mor

e ox

ygen

tha

n al

dehy

des.

)

In m

ost

case

s, r

anci

dity

is

due

to p

rodu

cts

of a

uto-

oxid

atio

n of

un

satu

rate

d

fatt

y ac

ids

by

prog

ress

ive

addi

tion

of

ox

ygen

to

lip

id

com

pone

nts.

A

utox

idat

ion

reac

tion

s ar

e ca

taly

sed

by p

rese

nce

of

proo

xida

nts

like

Cu, F

e, s

unlig

ht a

nd l

ipox

igen

ase

enzy

me.

The

shel

flif

e of

fat

s an

d oi

ls is

ass

umed

as

the

end

of t

he

indu

ctio

n pe

riod

, af

ter

whic

h th

e ra

te o

f fo

rmat

ion

of

oxid

atio

n pr

oduc

ts (

Pero

xide

s,O

xide

s, H

ydro

pero

xide

s)

incr

ease

s dr

asti

cally

. IP

is 1

.3 h

ours

for

C18

:3, 1

9 ho

urs

for

C18:

2, a

nd 8

2 ho

urs

for

C18:

1 at

40°

C.D

iffe

rent

te

sts

have

be

en

deve

lope

d to

m

easu

re

the

exte

nt o

f au

toxi

dati

on, b

ut e

ach

resp

ecti

ve t

est

is li

ke a

sn

apsh

ot a

t th

e ti

me

test

ed a

nd d

o no

t ne

cess

arily

sho

w th

e co

mpl

ete

pict

ure.

Qua

ntity

of

R

eact

ants

and

Prod

ucts

Qua

ntity

of

R

eact

ants

and

Prod

ucts

TIM

E

Indu

ctio

n pe

riod*

Seco

ndar

y ox

idat

ion

prod

ucts

Pero

xide

s

Prim

ary

oxid

atio

n pr

oduc

ts

*Ind

ucti

on

peri

od

is

the

leng

th

of

tim

e be

fore

de

tect

able

ra

ncid

ity,

af

ter

whic

h th

ere

is

an

acce

lera

ted

lipid

oxi

dati

on

AN

ALY

SIS

OF

FLA

VO

R Q

UA

LITY

& S

TAB

ILIT

Y O

F O

IL

Pero

xide

Val

ue

KI

C

H3

C

O

HH

I

CH

3

C

OK

OO

RO

OH

H

II 2

H

2O

RO

H

I 2

Na 2

S2O

3N

aI

Na 2

S4O

6

A.

B.

C.

+

+

+

+

+

+

+

2

2

2

Pero

xide

Val

ue =

m

l of N

a2S2

O3

×N

×10

00

(mill

iequ

ival

entp

erox

ide/

kg o

f sam

ple)

G

ram

s of O

il

Act

ive

Oxy

gen

Met

hod

(AO

M)

Det

erm

ined

the

time

requ

ired

to o

btai

n ce

rtain

pe

roxi

de v

alue

und

er sp

ecifi

c ex

perim

enta

l con

ditio

ns.

The

larg

er th

e A

OM

val

ue, t

he b

ette

r the

flav

or

stab

ility

of t

he o

il.

TBA

Tes

t

To d

eter

min

e th

e ra

ncid

ity d

egre

e of

mea

t or f

ish

prod

uct.

N

N

HS

OH

OH

C

CH

2

CO

HO

H

OH

OH

HS

N

N

OH

SH

N

N

CH

C

H

CHH

O

H2O

Col

ored

Pig

men

t

+

+2

Stor

age

stab

ility

(sh

elf-

life)

:Li

pids

var

y in

the

ir s

usce

ptib

ility

to

ranc

idit

y be

caus

e of

in

here

nt

prop

erti

es

like

unsa

tura

tion

, pr

esen

ce

of

anti

oxid

ants

, an

d so

me

exte

rnal

fac

tors

lik

e pr

oces

sing

an

d st

orag

e co

ndit

ions

.Acc

eler

ated

tes

ts a

re u

sed

for

pred

icti

ng f

utur

e be

havi

our.

For

exa

mpl

e, b

y ra

isin

g th

e te

mpe

ratu

re,

you

natu

rally

acce

lera

te

the

rate

of

de

teri

orat

ion.

At

the

end

of t

he in

duct

ion

peri

od,y

ou s

top

and

perf

orm

per

oxid

e de

term

inat

ion

anal

ysis

.En

d of

she

lf li

fe c

an b

e de

fine

d as

day

s at

20°

C to

rea

ch

P.V.

=20.

This

poi

nt i

s ge

nera

lly t

he e

nd o

f th

e in

duct

ion

peri

od,

and

the

end

of t

he t

est.

The

long

er t

he I

P, t

he

bett

er t

he s

tora

ge s

tabi

lity

and

the

long

er t

he e

xpec

ted

shel

flif

e.Fo

r th

is p

urpo

se m

any

diff

eren

t “a

ccel

erat

edsh

elf-

life”

met

hods

hav

e be

en d

evel

oped

:

-Sch

aal

Ove

nte

st:

Oil

is

kept

at

63

°C

till

ranc

idit

y is

de

tect

ed

or

till

the

end

of

incu

bati

on

peri

od(A

ssum

ptio

n:1

day

of

incu

bati

on a

t 63

°C c

orre

spon

ds t

o 10

day

s at

20

°C.)

-Act

ive

oxgy

enm

etho

d (A

OM

):O

2is

bub

bled

th

roug

h dr

ied

and

filt

ered

oil

to

furt

her

acce

lera

te a

utox

idat

ion

at 1

10°C

, thi

s is

car

ried

ou

t ti

ll PV

incr

ease

s to

20

for

lard

and

100

for

ve

geta

ble

oils

. Th

e in

duct

ion

peri

od

is

dete

rmin

ed b

y di

scon

tinu

ous

mea

sure

men

ts o

f pe

roxi

de v

alue

or

sens

ory

eval

uati

on o

f ra

ncis

od

or.

OXID

ATIV

E ST

ABI

LITY

INDEX

(OSI

)O

xida

tive

Stab

ility

Inde

x te

st, u

nlik

ete

sts s

uch

as p

erox

ide

valu

e w

hich

det

erm

ine

oil q

ualit

y on

ly a

t the

tim

e of

the

test

, ha

spre

dict

ive

valu

e, th

ereb

y al

low

ing

qual

ity d

ecis

ions

to

be m

ade

befo

re sp

oila

ge b

ecom

es a

pro

blem

. Th

e 0S

I is a

n in

valu

able

qua

lity

cont

rol t

ool f

or e

limin

atin

g in

ferio

r ing

redi

ents

(for

exa

mpl

e ha

zeln

uts f

or c

hoco

late

m

anuf

actu

re),

for m

easu

ring

and

com

parin

g th

e st

abili

ty o

f fin

ishe

d oi

l-fat

pro

duct

sas w

ell a

s lo

w m

oist

ure

snac

ks li

ke

pata

to c

hips

, for

eva

luat

ing

antio

xida

nt a

ctiv

ity, f

or

mon

itorin

g th

e de

terio

ratio

n of

coo

king

oils

with

use

, and

fo

r man

y ot

her a

pplic

atio

ns w

here

oxi

dativ

e ra

ncid

ity is

the

caus

e of

pro

duct

failu

re. B

ecau

se th

e O

SI h

as p

redi

ctiv

e va

lue,

pro

blem

s are

det

ecte

d an

d el

imin

ated

bef

ore

they

ca

use

reje

ctio

n, u

nlik

e ot

her t

ests

whi

ch m

erel

y in

dica

te th

at

prob

lem

s hav

e oc

curr

ed a

fter i

t is t

oo la

te to

cor

rect

them

.

-RA

NCI

MA

TM

etho

d:Ra

ncim

at v

alue

at

Tem

pera

ture

T°

is t

he n

umbe

r of

hou

rs,t

, bef

ore

the

oil r

each

es

the

end

of it

s in

duct

ion

peri

od.T

his

devi

ce p

erfo

rms

shel

f lif

e an

alys

is f

or a

t le

ast

2 pr

esel

ecte

d hi

gh

tem

pera

ture

s (i.

e. 1

00 °

and

130

° o

r 14

0°C)

.

For

this

pur

pose

, yo

u pu

t th

e sa

mpl

e in

the

dev

ice,

in

crea

se t

he t

empe

ratu

reto

the

pre

sele

cted

T1,

and

the

prod

ucts

of

oil o

xida

tion

und

er t

hese

con

diti

ons

are

colle

cted

by

dist

illat

ion

into

a w

ater

tra

p(a

spec

ial c

olle

ctin

g ve

ssel

con

tain

ing

deio

nize

d wa

ter)

. Th

e co

nduc

tivi

ty o

f wa

ter

is c

onti

nuou

sly

mea

sure

d an

d r

ecor

ded.

The

abr

upt

cha

nge

in c

ondu

ctiv

ity

mar

ks t

he e

nd o

f th

e in

duct

ion

peri

od,t

.

Ass

umpt

ions

:Q

10=

rate

at

(T+1

0) °

C/ r

ate

at T

°C

= sh

elfl

ife

at T

°C

/ sh

elfl

ife

at (T

+10)

°C

Grap

hs w

here

Co

nduc

tanc

e va

lue(

1/R)

is p

lott

ed

agai

nst

tim

e(t)

are

use

d fo

r sh

elfl

ife

pred

icti

on.

A p

lot

of lo

g t

(tim

e pe

riod

cor

resp

ondi

ng t

o en

d of

she

lf-

life)

ver

sus

T°C

thro

ugho

ut a

ran

ge o

f te

mpe

ratu

res

will

be

a st

raig

ht li

ne. T

he t

wo p

oint

s co

rres

pond

ing

to lo

gt1

and

log

t2 v

alue

s at

tem

pera

ture

s T

1an

d T 2

resp

ecti

vely

wi

ll

be p

lott

ed a

nd t

he t

wo p

oint

s so

con

nect

ed a

s to

in

terc

ept

both

x-a

xis

and

y-ax

is. T

he s

helf

life

at

a th

ird

tem

pera

ture

T 3

ca

n th

en

be

read

fr

om

grap

h by

ex

trap

olat

ion.

1/R

T(tim

e)

Log

t (ti

me)

Tem

pera

ture

°C

100°

140°

20°

For

refi

ned

edib

le v

eget

able

oils

, Ra

ncim

at v

alue

s ca

n be

use

d in

the

gen

eral

eq

uati

on o

f lin

e to

de

duce

exp

ecte

d sh

elf-

life:

D20

≅7.4

0 R

100-

7.9

(her

e, r

=0.9

33)

Whe

re D

20=d

ays

at 2

0°C

to re

ach

PV=2

0

And

R10

0=R

anci

mat

val

ue in

hou

rs a

t 100

°C

Phys

ical

Met

hods

-The

rmal

Met

hods

*Diff

eren

tial s

cann

ing

calo

rimet

er*T

herm

ogra

vim

etric

and

pres

sure

di

ffer

entia

l cal

orim

etric

met

hods

*Oxi

dativ

e st

abili

ty-le

ngth

of t

he

incu

batio

n pe

riod-

time

to m

ax w

eigh

t ga

in a

nd ra

te o

f wei

ght i

ncre

ase-

time

of m

ax e

ntha

lpy

chan

ge

Ther

mal

Met

hod-

Che

milu

min

esce

nce

*Det

erm

ine

the

cont

ent o

f ef

fect

iven

ess o

f ant

ioxi

dant

s in

oils

and

fats

*cor

rela

tes w

ith th

e pe

roxi

de

valu

e an

d an

isid

ine

valu

es a

nd

flavo

ur*e

mitt

ed d

urin

g pe

roxi

date

brea

kdow

n

Indi

cato

rs o

f Qua

lity

of H

eate

d O

ils *Det

erio

tive

chan

ges-

ther

mal

oxid

atio

n,hy

drol

ysis

,po

limer

izat

ion

*Dee

p fa

t fry

ing

pota

toes

with

fast

turn

over

in s

unflo

wer

oi

l-the

rmox

idat

ive

chan

ges

*A d

ram

atic

rise

in p

olar

com

poun

ds; M

ore

than

25-

27%

po

lar l

ipid

s sh

ould

be

reje

cted

*NIR

S-d

eter

min

e th

e su

m o

f dim

eran

d po

lym

er

trigl

ycer

ides

and

aci

d va

lue

to e

valu

ate

fryin

g oi

lC

hang

es in

die

lect

ric c

onst

ant

Tox

ic C

onta

min

ants

and

A

dulte

rant

s

-Con

tam

inat

ion

by w

eed

seed

s du

ring

harv

estin

g-S

tora

ge o

f saf

e oi

l in

unsa

fe

cont

aine

r

Con

tam

inat

ion

due

to F

aulty

Sto

rage

*Con

tam

inat

ion

of v

eget

able

oi

ls b

y m

iner

al o

ils-

dete

rmin

ing

the

sapo

nific

atio

nva

lues

of t

he lu

bric

atin

g oi

l sa

mpl

es*

Indi

ces o

f Adm

ixtu

res,

Ble

nds,

Con

tam

inan

ts a

nd

Adu

ltera

nts-

One

Fat

to

Ano

ther

*Sta

ble

carb

on is

otop

e ra

tio

anal

ysis

-est

ablis

hing

the

auth

entic

ity o

f oils

*Fou

rier t

rans

form

infra

red

spec

trosc

opy

for a

uthe

ntic

ity

of e

xtra

virg

in o

live

oil

Adm

ixtu

re o

f Veg

etab

le O

ils

with

Oth

er V

eget

able

Oils

Bio

synt

hesi

sof

Fat

-Spe

cies

spe

cific

-Det

ectio

n of

adu

ltera

tion

can

be b

ased

on

the

basi

s of

var

ious

con

stitu

ents

in

the

fat a

dmix

ture

Adm

ixtu

re o

f Veg

etab

le O

ils

with

Oth

er V

eget

able

Oils

-Fat

ty A

cid

Com

posi

tion

-Trig

lyce

ride

Ana

lysi

s-U

nsap

onifi

able

Frac

tion

of O

il*S

tero

l ana

lysi

s*T

ocop

hero

lana

lysi

s*P

heno

lics

and

Alco

hols

Fatty

Aci

d C

ompo

sitio

n*W

ithou

t bei

ng d

etec

ted

by th

e ro

utin

e ph

ysic

al a

nd c

hem

ical

cha

ract

eris

tics-

GLC

an

alys

is o

f fat

ty a

cids

pro

ve u

sefu

l-pr

esen

ce o

f so

ybea

n oi

ls in

oliv

e oi

l *I

OO

C fo

r oliv

e oi

l pur

ity-s

atur

ated

fatty

ac

id c

onte

nt a

t pos

ition

-2*P

ancr

eatic

lipa

se a

llow

s th

e di

ffere

ntat

ion

betw

een

virg

in a

nd e

ster

ified

oliv

e oi

ls-

Sat

urat

ed fa

tty a

cid

leve

l <1,

5% in

virg

in

oliv

e oi

l

Fatty

Aci

d C

ompo

sitio

n*C

ocon

ut o

il -d

eter

min

atio

n of

sap

onifi

catio

nan

d IV

-det

ect g

ross

and

sim

ple

adul

tera

tion

*Ana

lysi

s of

met

hyl e

ster

s of

fatty

aci

ds b

y G

C-

ratio

sof

cer

tain

fatty

aci

ds-d

etec

tion

of 5

%

rape

seed

oil

in p

eanu

t oil(

C22

-C16

)P

alm

ker

nel a

nd c

ocon

ut o

ils in

adu

ltera

ted

coco

a bu

ttera

s w

ell a

s ad

ditio

n of

coc

onut

or

palm

ker

nel o

il to

but

ter (

high

laur

ican

d m

yris

ticac

id c

onte

nt).

*Lin

olen

icac

id-d

etec

tion

of 5

% g

rape

seed

oil o

r <5

% s

unflo

wer

oil

in o

live

oil

Uns

apon

ifiab

leFr

actio

n of

Oils

*The

pre

senc

e or

abs

ence

of

certa

in in

divi

dual

uns

apon

ifiab

leco

mpo

nent

s ca

n he

lp in

es

tabl

ishi

ng th

e id

entit

y of

eac

h oi

l (a

uthe

ntic

ity o

r ge

nuin

enes

s)an

d al

so in

det

ectio

n of

adm

ixtu

re w

ith

anot

her o

il.

Uns

apon

ifiab

leFr

actio

n of

Oils

Ster

ol A

naly

sis

*Ana

lysi

s of

ste

rols

from

uns

apon

ifiab

lefra

ctio

n -a

dulte

ratio

n ol

ive

oil w

ith s

oybe

an o

il*S

tero

l com

posi

tion

obta

ined

by fr

actio

n of

the

unsa

poni

fiabl

em

atte

r is

char

acte

ristic

of v

irgin

ol

ive

oils

*The

ratio

of t

he c

onte

nt o

f β-s

itost

erol

to th

e su

m o

f (s

tigm

aste

rola

nd c

ampe

ster

ol)i

s 25

-30

for v

irgin

oliv

e oi

l.V

irgin

oliv

e oi

l has

β-s

itost

erol

cont

ent o

f 94%

or

hig

her

Uns

apon

ifiab

leFr

actio

n of

Oil

Toco

pher

olA

naly

sis

*Con

cent

ratio

n of

γ-to

coph

erol

sin

so

lven

t ext

ract

ed c

otto

nsee

d oi

ls

incr

ease

d on

con

tam

inat

ion

with

so

ybea

n oi

l*C

onta

min

atio

n of

oliv

e oi

l with

pe

anut

oil

-det

ecte

d fro

m

ingr

ease

dβ-

toco

pher

olle

vels

*Qua

ntita

tive

dete

rmin

atio

n of

δan

d γ-

toco

pher

ols

-a m

etho

d fo

r de

term

inin

g th

e ad

ulte

ratio

n of

ol

ive

oil w

ith le

ss e

x pen

sive

oils

Uns

apon

ifiab

leFr

actio

n of

Oil

Phen

olic

sand

Alc

ohol

s

*Rev

erse

pha

se H

PLC

with

UV

de

tect

ion

and

GC

-MS

-qua

ntifi

catio

n of

ph

enol

s*O

live

oil p

ress

cak