1. Omega-3: Ingredients and
Stability Monitoring
Dr. Steve Li, MD, M.Sc. (Clin. Pharm)
Director of Laboratory Operations

2. Outline
1. Omega-3 and its Product Development
2. Quality Control and Issues with Omega-3
Product Testing
3. Stability Study
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3. What is omega-3?
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4. Omega-3 fatty acids
They are polyunsaturated fatty acids
α-linolenic acid (18:3, n-3; ALA), eicosapentaenoic acid (20:5, n-3; EPA), and
docosahexaenoic acid (22:6, n-3; DHA)
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5. Essential fatty acids
ω-3 fatty acids:
– α-Linolenic acid or ALA (18:3n-3)
• ω-6 fatty acids:
– Linoleic acid or LA (18:2n-6)
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6. Fatty acid
synthesis in the
human body
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7. Why do we need it?
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8. Omega-3 FA and our health
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9. Omega-3 products
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10. Common omega-3 dosage forms
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11. The common active ingredients added in
omega-3
1. Minerals
• Calcium
• Selenium
• Chromiun
• Zinc
2. Vitamins
• Vitamin D
• Vitamin A
• Coenzyme Q10
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3. Antioxidants
• Vitamin E (alpha and
gamma tocopherols)
• Vitamin C and
derivatives (Ascobyl
palmatate)
4. Others (Antibiotics,
Hormones or Biologics)

12. Formulated products
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13. New product development
• Applications (Enteral, Parenteral and Topical)
• Taste and Appearance
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• Bioavailability
•Multiple health benefits
• Stability

14. Omega-3 products
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Eggs Yogurts Nutrition
Drinks
Meats
Cheese
Medicinal
Formulas
Bread Cosmetics

15. Now let’s see if we really do quality
control for the products
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16. 3 QbD things useful for omega-3-like
products
1. Can a product be easily tested?
a) Is there a reliable method or compendial method
for testing a product?
b) Has a method been verified for a product?
2. Is the formulation compatible?
3. Is a product stable in storage?
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17. Challenges of quality control methods for
testing omega-3 products
• Oil Based Matrix---- Solubility and
Homogeneity
• Continuous Oxidation Process----Stability
• Complexity of Formulation----Interfering
Additives or
Ingredients for testing
• Less Specific----Less Supportive.
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18. Test methods for omega-3 fatty acids
• Fatty Acid Oxidation
• Release Testing of API and Products
• Microbiological Tests
• Heavy Metal Tests
• Pesticides Tests
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19. Oxidation is the main degradation
pathway of omega-3
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Oxidation Heat & Light
Degradation

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

21. Acceptable limits for oxidative stability
parameters in marine oils
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(Recommended Reporting Threshold)
________________________________________
Acid Value 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 Anisidine Value TAVTM 70% of Anisidine Remaining
______________________________________________________

22. Acid Value Test
Titration Process and Color Change
Steve Lianghong Li, NDI ADRL dba Diteba

23. Peroxide value test
Titration Process and Color Change
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24. P-Anisidine value test
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25. p-Anisidine HPLC-UV method
0.005
0.004
0.003
0.002
0.001
1.00 2.00 3.00 4.00 5.00 6.00 7.00 8.00 9.00 10.00
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Typical Chromatogram of QL
(S/N=15, 3.162 μg/mL)
3.305
p-Anisidine - 4.855
Channel @ W2996 PDA 254.0 nm at 1.2
AU
0.000
Minutes

26. Quantitative assay method for omega-3
• Most common method is GC-FID or GC-MS
• LCMS can be an good alternative.
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27. Stability Studies
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28. Unique features of omega-3 containing
products and stability testing
• Most of them are softgel capsules, oil liquid or oil
based suspension
• Most common degradation is oxidation and the
degradation impurities are mixtures of multiple
compounds.
• It is extremely difficult to have a stability indicating
method for Omega-3 fatty acids since the degradation
products are variable and difficult to be easily identified
and quantified using simply analytical methods.
• Most of current methods for testing oxidation products
are not specific and directly quantitative for the
degradation products.
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29. Challenges in quality control and stability
study of omega-3 products
• Oil Based Matrix-Homogeneity
• Unstable Due to Continuous Oxidation Process
• Need More Efficient and Specific QC Analytical
Methods
• Formulated Product with Interfering Additives
or Ingredients for the Current Test Methods
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30. General storage conditions in stability
studies
• Storage ICH
Walk-in and Reach-in Chambers
• Storage Conditions
25°C/60% RH
30°C/65% RH
40°C/75% RH
Photostability
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31. Specifications in stability study
• Stability studies should include testing of those attributes of
the API that are susceptible to change during storage and
are likely to influence quality, safety and/or efficacy.
• The testing should cover, as appropriate, the physical,
chemical, biological and microbiological attributes. A guide
as to the potential attributes to be tested in the stability
studies.
• Validated stability-indicating analytical procedures should
be applied.
• Whether and to what extent replication should be
performed will depend on the results from validation
studies
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32. Stability and testing frequency
• For APIs with a proposed re-test period or shelf-life of at
least 12 months, the frequency of testing at the long-term
storage condition should normally be every three months
over the first year, every six months over the second year,
and annually thereafter throughout the proposed re-test
period or shelf-life.
• At the accelerated storage condition , a minimum of three
time points, including 0, 3, and 6 month.
• When testing at the intermediate storage condition a
minimum of four time points, including the initial and final
time points (e.g. 0, 6, 9 and 12 months), from a 12-month
study is recommended.
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33. Typical example of stability program
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34. Typical stability program
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35. Compatibility studies (3 month)
Formula 1
% LC Oxidation
Total EPA+DHA CoQ 10
Time
(Month)
Time
(Month)
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p-Anisidine
Value
TAV
(%)
Peroxide
Value
(mEq/Kg)
Acid
Value
(mg
KOH/g)
% Total
Impurities
0 98.5 102 120 85 6 1 1.00
3 96.8 96 160 76 10 1.8 1.52
Formula 5
% LC Oxidation
Total EPA+DHA 96Micro Extract A
p-Anisidine
Value
TAV
(%)
Peroxide
Value
(mEq/Kg)
Acid
Value
(mg
KOH/g)
% Total
Impurities
0 98.5 94 12 85 4 0.5 2.50
3 96.8 72 160 76 10 1.8 8.00

36. Example of stability summary
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% LC
Month Batch 1 Batch 2
0 98.5 98.5
3 96.8 96
6 94.7 94
9 93.3 92.5
12 92.3 86
18 92.9 80
24 91.6 76
36 90.3 75

37. Fancy term but common logic
Quality by Design (QbD)
1. Analytical Methods
2. Compatibility Study (3 Month)
3. Specifications
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38. QbD principles
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Desired
State of
Product
ICH Q8-
Pharm.Dev
elopment
ICH Q10
Quality
System
ICH Q9-Quality
Risk
Management

39. Fate of drug product and philosophy
• Only Very Small % Drug Products Get FDA Approval
Product
Philosophy
Fate of
Product
Application
Confirmation
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Formula
Quality
Control
Tox
Product
Philosophy
Fate of
Product
Formula
QbD
Quality
Control
Application
Confirmation
Tox

40. Thank you!
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Dr. Steve Li, MD, M.Sc. (Clin. Pharm)
Director of Laboratory Operations
stevel@diteba.com
Connect with Diteba
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