Effect of oxidized fat in food on lipoprotein oxidation

9.5.2012 | Jukka-Pekka Suomela

Department of Biochemistry and Food Chemistry

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BackgroundFat oxidation

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(autoxidation)

This proton ripped off from anotherfatty acid molecule

BackgroundFat oxidation

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BackgroundFat oxidation

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Breakdown of hydroperoxides

BackgroundFat oxidation

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Photosensitizedoxidation of linoleicacid

Autoxidation oflinoleic acid

(Buettner, Molecular targets of photosensitization)

In addition, enzymatic oxidation is possible inbiological systems…

BackgroundFat oxidation

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Background“Basic” indicators of fat oxidation

peroxide value (PV, POV) Fat oxidation gives birth to peroxides that are

measured with a method based on the ability of peroxides to release iodine from potassium jodide

Titration of the relased iodine with Na2S2O3-liuoksella

p-anisidine value (p-AV) Measures the amount of secundary oxidation products

(aldehydes, ketones) in oil spectrophotometric determination at 350 nm

Fats get rancid also hydrolytically!

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BackgroundOxidation products from dietary fats to the body –

lipoproteins and atherosclerosis

Witztum, J.L. Lancet 344: 793-795

Oxidized lipoproteins seem to increase the risk of atherosclerosis LDL Chylomicrons, VLDL, and their remnants?

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BackgroundClassification of lipoproteins

chylomicrons formed in small intestine

very low density lipoproteins (VLDL) formed in liver

intermediate density lipoproteins (IDL) formed from VLDL

low density lipoproteins (LDL) formed from IDL

high density lipoproteins (HDL) formed in liver and small intestine

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Durrington, P.N. Hyperlipidaemia: Diagnosis and Management. 2nd edition.

London: Butterworth-Heinemann; 1995.

BackgroundClassification of lipoproteins

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BackgroundMetabolism of lipoproteins

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BackgroundFormation of foam cells

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BackgroundAtherosclerotic plaque

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BackgroundLipoproteins transport oxidized lipids

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BackgroundFood and oxidized lipoproteins

Staprans, I. et al. Arterioscler Thromb 14: 1900-1905

In various studies, dietary oxidized lipids have increased the oxidation of chylomicrons and VLDL

food small intestine lipoproteins other tissues… What? Where? Why?

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Aim of the research

1. To study the effect of oxidized dietary oil on the amount of oxidized lipids in porcine lipoproteins

2. To identify the molecular structures of oxidized triacylglycerols (TAGs) in porcine chylomicrons and VLDL

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Oxidation of fatty acids within glycerolipids

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Study protocols

STUDY 1 3 groups of 3 pigs; 14%

sunflower seed oil in the feed Peroxide value of the oil:

group 1: 1 meq O2/kg oil

group 2: 84 meq O2/kg oil

group 3: 223 meq O2/kg oil

STUDY 2 2 groups of 10 pigs; 16%

sunflower seed oil in the feed Peroxide value of the oil:

group 1: 1 meq O2/kg oil

group 2: 190 meq O2/kg oil

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Methods

1. Determination of the oxidation level of pig lipoproteins BDC (baseline diene conjugation) spectrophotometrically

2. Identification of the molecular structures of oxidized TAGs in chylomicrons and VLDL: Separation of lipid classed by TLC (thin layer chromatography) Further separation by RP-HPLC (reversed-phase liquid

chromatography) Detectors:

ESI-MS ELSD UV (DNPH derivatives of core aldehydes)

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MethodsIdentification of molecular structures, TLC

TAGOxidizedTAG

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(MeOH / i-Pr) (80 / 20, by vol)

0,85 ml / min20 min

(MeOH / i-Pr) (20 / 80, by vol)

SOLVENT GRADIENT:

Discovery HS C18 5 m, 250 mm x 4.6 mm i.d.

COLUMN:

MethodsIdentification of molecular structures, HPLC

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HPLC

130 l/min850 l/min

720 l/min

UV/ELSD

ESI MS

MethodsIdentification of molecular structures, HPLC

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Methods

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ResultsStudy 1

0

5

10

15

Time point 1 Time point 2

BD

C (

mol

/L)

Group 1

Group 2

Group 3

Chylomicrons

0

1

2

3

4

5

6

Time point 1 Time point 2

BD

C (

mol

/L)

Group 1

Group 2

Group 3

VLDL

LDL

0

5

10

15

20

25

Time point 1 Time point 2

BD

C (

mol

/L)

Group 1

Group 2

Group 3

a

b

Group 1: 1 meq O2/kg oilGroup 2: 84 meq O2/kg oilGroup 3: 223 meq O2/kg oil

Time point 1: 3 hr after mealTime point 2: 4 hr after meal

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ResultsStudy 1

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Estimated proportions of oxidized TAGs of the total lipids of the test oils and chylomicronsa

Group 1b Group 3c

Oil 0.2 Oil 6.2

Time point 3 h Time point 3 h Pig 1 0.0 Pig 7 0.4Pig 2 0.1 Pig 8 0.4Pig 3 0.2 Pig 9 1.9

Time point 4 h Time point 4 h Pig 1 0.0 Pig 7 0.6 Pig 2 0.3 Pig 8 1.1 Pig 3 0.3 Pig 9 1.2

aResults as g/100 g total lipids.bPeroxide value of the oil used in feed: 1 meq O2/kg.cPeroxide value of the oil used in feed: 223 meq O2/kg.

ResultsStudy 1

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Typically only one fatty acid residue of a TAG molecule was oxidized in the identified molecules

Among oxidized TAG structures were: hydroxides epoxides ketones aldehydes (9:0, 12:1)

Hydroperoxides were not found

ResultsStudy 1

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0

5

10

15

20

25

30

35

Time point 1 Time point 2

BD

C (

mo/

L)

Group 1

Group 2a

b

Chylomicrons

0

5

10

15

20

25

30

35

Time point 1 Time point 2

BD

C (

mol

/L)

Group 1

Group 2

VLDL

0

5

10

15

20

25

30

35

Time point 1 Time point 2

BD

C (

mol/L)

Group 1

Group 2

LDL

Group 1: 1 meq O2/kg oilGroup 2: 190 meq O2/kg oil

0

10

20

30

40

50

60

70

80

Time point 1 Time point 2

BD

C (

mol

/L)

Group 1

Group 2

Plasma

a

b

ResultsStudy 2

Time point 1: 3 hr after mealTime point 2: 4 hr after meal

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Estimated proportions of oxidized TAGs of the total lipids of the test oils, chylomicrons, and VLDLa

Group 1b Group 2c

Oil 0.0 Oil 6.4

Chylomicrons

Time point 3 h 0,8 ± 0,7a Time point 3 h 2,0 ± 0,6b

Time point 4 h 0,8 ± 0,9a Time point 4 h 2,4 ± 0,7b

VLDLTime point 3 h 0,1 ± 0,1 Time point 3 h 0,2 ± 0,2Time point 4 h 0,1 ± 0,1a Time point 4 h 0,2 ± 0,1b

aResults as g/100 g total lipids.bPeroxide value of the oil used in feed: 1 meq O2/kg.cPeroxide value of the oil used in feed: 190 meq O2/kg.

ResultsStudy 2

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ResultsStudy 2

Typically only one fatty acid residue of a TAG molecule was oxidized in the identified molecules

Among oxidized TAG structures were: hydroxides epoxides ketones aldehydes (9:0, 12:1)

Hydroperoxides were not found

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Conclusions

Diet containing oxidized sunflower oil increased lipid oxidation in lipoproteins

Oxidized fats are partly transported to the circulation Effect of vitamin E?

Several oxidized molecular structures derived from TAGs were found in chylomicrons and VLDL Method was not optimized to extensively oxidized

structures

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Literature

Suomela, J.-P., Ahotupa, M., Sjövall, O., Kurvinen, J.-P., & Kallio, H. (2004) Diet and lipoprotein oxidation: analysis of oxidized triacylglycerols in pig lipoproteins. Lipids 39: 639–647, DOI: 10.1007/s11745-004-1277-4

Suomela, J.-P., Ahotupa, M., and Kallio, H. (2005) Triacylglycerol oxidation in pig lipoproteins after a diet rich in oxidized sunflower seed oil. Lipids 40: 437–444, DOI: 10.1007/s11745-005-1402-4

Ahotupa, M., Suomela, J.-P., Vuorimaa, T., and Vasankari, T. (2010) Lipoprotein-specific transport of circulating lipid peroxides. Ann Med 42: 521–529, DOI: 10.3109/07853890.2010.510932

Tarvainen, M., Phuphusit, A., Suomela J.-P., Kuksis, A., and Kallio H. (2012) Effects of antioxidants on rapeseed oil oxidation in an artificial digestion model analyzed by UHPLC−ESI−MS. J Agric Food Chem, DOI: 10.1021/jf2050944. E-pub ahead of print.

Kanner, J., Gorelik, S., Sirota, R., & Kohen, R. (2012) Protection by polyphenols of postprandial human plasma lipid oxidative modification: the stomach as a bioreactor. J Agric Food Chem, DOI: 10.1021/jf300193g. E-pub ahead of print.