A Presentation made as a part of my course on "Nutraceuticals & Functional Foods" (Bachelor's study)

1. Anas Ejaz Shaikh
13FET1006
Fatty Acids

2. • ω-3 Fatty Acids are PUFAs with double bond at
the third carbon from the end of the carbon chain.
• Three ω-3 Fatty acids important in Human
physiology-
1. Alpha Linolenic Acid (ALA
2. Eicosapentaenoic Acid (EPA)
3. Docosahexaenoic Acid (DHA)
• Known as essential fatty acids (EFAs) as the body
cannot make these fatty acids on its own. Must be
obtained from food.
• Sources include foods such as cold-water fish
including tuna, salmon, and mackerel. Other
important omega 3 fatty acids are found in dark
green leafy vegetables, flaxseed oils, and certain
vegetable oils.
What are ω-3 Fatty Acids? & Why ω-3
Fatty Acids ?

4. Structure of
ω-3 fatty
acids

5. Sources

9. Roles of Omega 3 and Omega 6 Fatty
Acids
• Essential Fatty Acids (not easily
synthesized by the body)
1) Normal Brain function
2) Growth and Development
3) Bone Health
4) Stimulation of Skin and Hair
Growth
5) Regulation of Metabolism
6) Maintenance of Reproductive
processes
Both omega-6 and omega-3 fatty acids can be converted into three different types
of active molecules called prostaglandins.
These prostaglandins, PGE1, PGE2 and PGE 3, are site specific.

10. • Metabolic Pathway :
• Omega 3-
• Anti-inflammatory
• -Thus,
Prevention of Cancer,
Diabetes mellitus and
Cardiovascular
Diseases
• Omega 6-
• Pro-inflammatory /
Anti-inflammatory
• - Depending
on omega 3 : omega 6
ratio

11. Need for Balanced Ratio of ω-6 : ω-3
• Evolution of Humans1 : 1
• Scenario ofWestern diets15-17 : 1
• 70% Reduction in mortality in
secondary CHD prevention
4 : 1
• Reduced rectal cell proliferation
in patients with colorectal cancer2.5 : 1
• Suppressed Inflammation in
patients with rheumatoid
arthritis
2-3 : 1
• Beneficial effect on Asthama5 : 1

12. Natural Ratios of ω 6: ω3
• Most Plant oils – no ω-3
long term ingestion-
chronic diseases
• Animal fats (except
Chicken fat)-Balanced ω-
6: ω-3 ratio-but high in
saturated fat
• Canola oil-Balanced ω-6:
ω-3 ratio; rich in MUFAs
• Olive oil-Moderately
balanced ω-6: ω-3 ratio;
MUFAs; Anti-oxidants;
Squalene (Anti-Cancer)
• Fish Oils and Flaxseed Oil
: Best source of ω-3

13. Fatty acids in nuts

14. Reducing ω 6 and Increasing ω 3

15. FDAAnnounces Qualified
Health Claims for Omega-3
Fatty Acids
• The Food and Drug Administration (FDA) today announced the
availability of a qualified health claim for reduced risk of
coronary heart disease (CHD) on conventional foods that contain
eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA)
omega-3 fatty acids.

16. Health Benefits of ω-3 fatty acids

17. ω -3 fatty acids in Eye Health :
Prevention of Dry Eye Syndrome

18. • Good health depends on omega-6 gamma linolenic acid, a
downstream metabolite of linoleic acid.This compound is a
necessary component in the downstream metabolism of
omega-6 fatty acid to PGE1, which is associated with healthy
mucosal tissue and healthy tear film.

19. Inflammation leads to Diabetes and CVD

20. The remnant lipoprotein (RLP), produced from the triacylglycerol-rich chylomicrons and VLDL, exerts potent
pro-atherogenic effects.
The inner layer of the blood vessel wall (the endothelium) becomes infiltrated by inflammatory cells,
particularly macrophages andT-cells.
Actually, cholesterol in the sub-endothelial space is ingested by macrophages resulting in foam cells
In acute heart disease, macrophages and T cells become activated and secrete a number of inflammatory
cytokines and enzymes that disintegrate the fibrous cap, resulting in the clotting of blood.

21. Inflammatory alterations in adipose tissue
become evident by accumulation of
inflammatory cells such as macrophages
and T-lymphocytes, activation of
inflammatory signaling pathways and
production of inflammatory cytokines and
chemokines, collectively called adipokines.
Chemokines attract inflammatory cells,
while inflammatory cytokines interfere
with insulin action in metabolic cells by
targeting specific molecules.
Molecules that promote atrraction of
inflammatory cells or induce inflammatory
signaling pathways hence elicit insulin
resistance, the basis of type 2 diabetes.
Adipose tissue is not merely a storage
organ, but besides adipocytes (fat cells)
adipose tissue comprises a variety of cell
types including immune cells. The
picture shows immune cells (brown)
invading adipose tissue in obesity
Inflammation related to Type 2 Diabetes

23. Omega-3 fatty acids enhance synaptic transmission via the activation of energy-
generating metabolic pathways and by maintaining membrane ionic permeability

24. Metabolic Fate of fatty acids

25. Free fatty acids-most bioavailable as they surpass enzymatic hydrolysis
Some products like KrillOil claim greater bioavailability due to phospholipid presence
Some authors claim slight higher bioavailabilty ofTG than EE
Bioavailability of Different forms of omega 3 fatty acids

26. Phospholipids are more bioavailable than TG

27. Oxidation is the main degradation
pathway of omega-3
www.diteba.com
Heat & LightOxidation
Degradation
Stability of omega 3 fatty acids

28. Omega-3 fatty acid oxidation tests
www.diteba.com
• Acid Value: Titration method,
colorimetric
• p-Anisidine Value: UV 350 nm
• True Anisidine Value: HPLC UV
• Peroxide Value: Titration method,
colorimetric

29. Acceptable limits for oxidative stability
parameters in oils
www.diteba.com
(Recommended ReportingThreshold)
________________________________________
AcidValue NMT 3 mg KOH/Kg
Peroxide value (PV) NMT 5 mEq/Kg
Anisidine value (AV) NMT 20
TOTOX value NMT 26 (2 x PV + AV )
True AnisidineValueTAVTM 70% of Anisidine Remaining
______________________________________________________

30. Acid Value Test
• An increment in the amount of
FFA in a sample of oil or fat
indicates hydrolysis of
triglycerides.
• Such reaction occurs by the
action of lipase enzyme and it
is and indicator of inadequate
processing and storage
conditions (i.e., high
temperature and relative
humidity, tissue damage).

31. Peroxide value test
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Titration Process and Color Change
The fat or oil is dissolved in a chloroform-acetic acid mixture and subjected to an
excess of iodide via a saturated solution of potassium iodide.
The peroxides present oxidize the iodide to iodine and the iodine is then titrated to a
colorimetric endpoint using sodium thiosulfate with starch as an indicator.
The amount of iodine produced is directly proportional to the peroxide value.

32. p-Anisidine value test
Aldehydes, derived from the secondary
oxidation of fat matrix, react with the p-
anisidine determining a variation in the
absorbance, measured at 366 nm.
Anisidine value test is used to assess the
secondary oxidation of oil or fat, which is
mainly imputable to aldehydes and ketones,
and is therefore able to tell the oxidation
"history" of an oil or a fat.
Furthermore, AnV analysis on oil is an
indicator of excessive oil deterioration in
deep frying process.

33. p-Anisidine HPLC-UV method
www.diteba.com
Typical Chromatogram of QL
(S/N=15, 3.162 µg/mL)
Channel @ W2996 PDA 254.0 nm at 1.2
3.305
p-Anisidine-4.855
AU
0.000
0.001
0.002
0.003
0.004
0.005
Minutes
1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00

34. Quantitative assay method for omega-3
www.diteba.com
• Most common method is GC-FID or GC-MS
• LCMS can be an good alternative.

35. Typical example of stability program

36. • Extraction Methods for Obtaining Omega-3 Supplements
• Pressing The majority of oils come from cold water fish (sardine, anchovies). Use of
high temperatures (85-95 C) denatures the proteins of the fish.
• Protein aqueous phase separated from oil phase.
• Oil phase bleached and purified by addition of amorphous silica and then deodorized.
This process gives fish oil.
• Fractionation & Purification of Omega 3 Fatty Acids-
1. Cryoconcentration: Omega-3 fatty acid enrichment by crystallization at low
temperature. Lowering the temperature to less than -50 C in the presence of acetone.
2. Molecular distillation: First esterification is done using ethanol to form fatty acid
ethyl esters which can be separated according to differences in their boiling points
and molecular weights.
3. Chromatographic techniques: Based on separation of fatty acids according to the
length of their carbon chains or their degree of unsaturation permits high purification
of omega-3’s at levels surpassing 90%
Extraction of omega-3 fatty acids

37. • Molecular distillation is vacuum distillation below the pressure of
0.01 torr (1.3 Pa)
1: Still pot with stirrer bar/anti-bumping
granules
2: Cold finger – bent to direct condensate
3: Cooling water out
4:cooling water in
5:Vacuum/gas inlet
6:Distillate flask/distillate.

38. Supercritical Extraction

39. Benefits of Supercritical Extraction
1) Supercritical CO2 is used as a solvent to extract Lipophilic
compounds from natural herbs.
2) These extracts are concentrated as high as 250 times as
compared to the raw herb.Thus a small quantity in any product
is enough, leading to cost effectiveness as compared to other
products from different extraction techniques.
3) This technology is environment friendly with minimum carbon
footprint and CO2 is recycled as much as 95% in the system.

40. Liquid Chromatography

42. Nutrient-Nutrient Interaction
• Presence of excess fat in diet decreases Thiamine requirement.
• Similarly essential fatty acids can in a low Pyridoxine diet offers protection
against Pyridoxine deficiency.
• Vitamin E, the primary fat soluble antioxidant protects omega-3 fats from
oxidation
• The conversion of alpha-linolenic acid to EPA and DHA involves a series of
chemical reactions catalyzed by the enzymes like delta-6 desaturase &
delta-5 desaturase.
• To increase the activity of your desaturase enzymes, be sure that your diet
includes a sufficient amount of vitamin B6, vitamin B3, vitamin C,
magnesium and zinc.

44. Deficiency of Omega-3 fatty acids
• The following are associated with not getting enough omega-3:
• Fatigue
• Immune weakness
• Heart problems
• Mood swings or depression
• Poor memory
• Hair loss, dry skin or eczema
• Reproductive problems
• Poor circulation

45. Omega 3 Fatty acids Side Effects &
Toxicity..
• The fact of the matter is that when it comes to omega 3 side
effects and precautions, it really depends on the source of
the supplement.
• Today, the most popular omega 3 supplements come from
either fish oil, flaxseed oil, or cod liver oil so we will discuss
them separately.

46. Fish Oil Side Effects and
Precautions
• It appears that fish oil harbors possible side effects such
as belching, nausea, diarrhea, bloating, and intestinal gas.
• Fishy body odor and/or "fish breath," including burping a "fishy"
smell.
• Blood-thinning effect
• Beware
• : anticoagulant medications
• : bleeding disorder
• Test for contaminants like heavy metals

47. Cod Liver Oil Side Effects and
Precautions
• Cod liver oil contains both vitamin A and D
(Hypervitaminosis)
• Blood thinning medications such as Coumadin or over-the-
counter blood thinners such as aspirin, should consult with a
physician before taking any supplements.
• Test for contaminants like heavy metals

48. Flaxseed Oil Side Effects and
Precautions
• Appears to be safe
• Mild gastrointestinal symptoms such as diarrhea.
• People who suffer from gallbladder problems may
experience trouble digesting flax seed oil, which can be
resolved if you start with very small amounts (1 teaspoon)
and build up the dosage of flax seed oil over time.

49. 49
How to increase Omega-3 intake?
• Eat more fatty fish & nuts
• Take a supplement
• Eat fortified foods

51. Capsules
1. Low Potency - 300 mg
EPA+DHA/g =(Typical drug
store capsules)
2. High Potency - 500-700 mg
EPA+DHA/g (CardioTabs,
Triomega, OmegaRx)
3. Pharmaceutical – 850 mg
EPA+DHA/g (Omacor®,
Reliant Pharmaceuticals)
Supplements Available
Rs.1130/- for
60 capsules.
About
Rs.19/capsuleEPA and DHA are fatty acids and hence thermally stable up to
250 C, but the major problem is OXIDATIVE STABILITY.
Therefore, anti-oxidants are added to prevent its oxidation.
Most often tocopherol is used as an antioxidant.

53. Gel Technology by Agel

55. References
• http://www.fda.gov/
• www.consumerhealthdigest.com
• www.gbhealthwatch.com
• www.cholesterolbattle.com
• www.healio.com
• http://ako3.d393.dinaserver.com/en/krill-oil-16
• Amate L, Gil A, Ramírez M. Feeding infant piglets formula with long-chain
polyunsaturated fatty acids as triacylglycerols or phospholipids influences the
distribution of these fatty acids in plasma lipoprotein fractions. J Nutr. 2001:
131 (4); 1250 – 5.
• www.bioinfo.org.cn
• t3-cdl-cmit.meduniwien.ac.at
• www.nutritional-supplement-educational-centre.com/omega-3-side-
effects.html