1. Moisture content evaluation for highly hydrophobic natural compound mixtures / cyclodextrin complexes by thermal methods
Daniel I. Hădărugă, Nicoleta G. Hădărugă, Geza N. Bandur
Polytechnic University of Timişoara, Romania
Presented at “The 10th International Conference on Water in Food”, Prague, Czech Republic, September 19-21, 2018

2. OUTLINE Introduction:
Cyclodextrins & natural hydrophobic mixtures (oil)
Objective of the study
Methods, Results and Discussion:
Oil separation & Cyclodextrin complexation
Molecular modeling & Docking of highly hydrophobic compounds and cyclodextrins
GC-MS analysis & Thermal analysis (TG-DTG & DSC)
Principal Component Analysis (PCA)
Conclusion

3. Introduction: Cyclodextrins
Cyclodextrins are cyclic oligosaccharides obtained by enzymatic synthesis from starch.
They are
“GENERALLY REGARDED AS
SAFE (GRAS)”
and used in food and
pharmaceutical products.
Cyclodextrins can be used for:
* WATER-SOLUBILISATION
(apparent)
* ENHANCING BIOAVAILABILITY
* PROTECTION AGAINST
DEGRADATION
* CONTROLLED RELEASE
of various bioactive compounds
(GUESTS)

4. The mechanism of cyclodextrin complexation
Introduction:
Cyclodextrins
assumed to be
“strongly retained”
water molecules
assumed to be
“surface”
water molecules
Water as solvent
“bulk solvent”
The mechanism of cyclodextrin complexation
Ref. on water content of cyclodextrins (KFT,): Journal of Inclusion Phenomena and Macrocyclic Chemistry 2013, 75 (3-4),

5. Highly hydrophobic guest
Introduction:
natural hydrophobic mixtures (oil)
Triolein (blue) and DHA based triglyceride (red)
Very low stability!
FATTY ACID TRIGLYCERIDES
Monopalmitin
FATTY ACID MONOGLYCERIDE
EICOSAPENTAENOIC ACID (EPA)
Very low stability!
FREE FATTY ACID
Highly hydrophobic guest
compound mixtures
Ref. on thermal and oxidative stability of CD/linoleic acid complex: Hădărugă et al., Food Chemistry 2006, 99(3),

6. Objective of the study
The overall influence of the fatty acid profile of various highly hydrophobic natural compound mixtures (oily fruits, vegetable and fish oils) on the cyclodextrin complexation, through the hydration water/moisture content and behavior by thermal analyses

7. Methods, Results and Discussion:
Oil separation & Cyclodextrin complexation
Beans’ lipid fractions
(solid-liquid extraction) or
Walnuts and
common hazel oils
(pressing method)
Fish oils
(muscle, boiling-pressing method)
Cyclodextrin
complexation
Kneading
(or co-crystallization)
Drying & Grinding
Refs. on CD/fish oil complex:
Hădărugă, et al. Food Chemistry 2017, 236, 49-58
Hădărugă, et al. LWT – Food Science and Technology 2016, 68,
Hădărugă, et al. Beilstein Journal of Organic Chemistry 2016, 12,

8. Methods, Results and Discussion:
Molecular modeling & Docking of highly hydrophobic compounds and cyclodextrins
β-Cyclodextrin / Palmitic acid complex
(1:1 in vacuum)
(optimized by MM+ force field)
E(int) = 23 kcal/mol
β-Cyclodextrin / EPA complex
(1:1 in vacuum)
(optimized by MM+ force field)
E(int) = 19 kcal/mol

9. Methods, Results and Discussion:
Molecular modeling & Docking of highly hydrophobic compounds and cyclodextrins
Theoretical representation of the molecular encapsulation process of the Atlantic salmon oil glycerides (e.g. EPA triglyceride) and β-cyclodextrin

10. Methods, Results and Discussion:
Molecular modeling & Docking of highly hydrophobic compounds and cyclodextrins
3 x β-Cyclodextrin / Palmitin complex
(3:1 in vacuum)
(optimized by MM+ force field)
E(int) = ~90 kcal/mol
3 x β-Cyclodextrin / EPA triglyceride complex
(3:1 in vacuum)
(optimized by MM+ force field)
E(int) = ~90 kcal/mol
Ref. on molecular modeling and docking of FAs, their glycerides and bioconjugates:
Hădărugă et al., Journal of Agroalimentary Processes and Technologies 2017, 23(1), 5-12

11. Methods, Results and Discussion:
GC-MS analysis & Thermal analysis (TG-DTG & DSC)
(Common hazel oil and its β-cyclodextrin complex)
Retention index (RI)
Area, %
(H)
Fatty acid derivative
1908
0.70
Palmitoleic acid, methyl ester
1931
9.72
Palmitic acid, methyl ester
2092
12.09
Linoleic acid, methyl ester
2103
62.89
Oleic acid, methyl ester
2124
2.68
Stearic acid, methyl ester
2401
4.72
Linolenic acid, methyl ester / Linolenin, 1-mono-
12.4
SFA
63.6
MUFA
16.8
PUFA (especially omega-6)
GC-MS analysis of common hazel oil (derivatized to methyl esters)

12. Methods, Results and Discussion:
GC-MS analysis & Thermal analysis (TG-DTG & DSC)
(Common hazel oil and its β-cyclodextrin complex)
TG-DTG (top, 1:1) and DSC (right, 3:1) analyses of β-cyclodextrin / common hazel oil complexes at 1:1 molar ratio
Ref. on TG & DSC review:
Hădărugă et al., Springer Nature, 2018, pp

13. Methods, Results and Discussion:
GC-MS analysis & Thermal analysis (TG-DTG & DSC)
(Beans’ lipids and their β-cyclodextrin complex)
See: Poster No 6
GC-MS analysis of beans lipid fractions (derivatized to methyl esters)
Retention index (RI)
Area, %
(BN_SW)
(BN_NE)
Fatty acid derivative
1907
0.43 ± 0.02
0.47 ± 0.08
Palmitoleic acid, methyl ester
1934
17.36 ± 0.41
13.9 ± 1.8
Palmitic acid, methyl ester
2098
35.64 ± 1.24
48.61 ± 9.21
Linoleic acid, methyl ester
2105
31.63 ± 1.43
25.27 ± 5.59
Oleic acid, methyl ester
2124
2.46 ± 0.1
3.23 ± 0.14
Stearic acid, methyl ester
2405
3.23 ± 0.2
0.95 ± 0.99
Linolenin, 1-mono- (omega-3)
23.1 ± 0.48
20.22 ± 1.71
SFA
34 ± 1.64
27.03 ± 6.57
MUFA
40.48 ± 0.87
50.91 ± 7.83
PUFA (especially omega-6)

14. Methods, Results and Discussion:
GC-MS analysis & Thermal analysis (TG-DTG & DSC)
(Beans’ lipids and their β-cyclodextrin complex)
See: Poster No 6
TG-DTG (top) and DSC (right) analyses of β-cyclodextrin / beans’ lipids complexes at 1:1 molar ratio

15. Methods, Results and Discussion:
GC-MS analysis & Thermal analysis (TG-DTG & DSC)
(Asp oil and its β-cyclodextrin complex)
See: Poster No 14
Retention index (RI)
Area, %
(Asp oil)
Fatty acid derivative
1885
13.96 ± 0.40
Palmitoleic acid, methyl ester
1907
14.44 ± 0.17
Palmitic acid, methyl ester
2067
5.58 ± 0.44
Linoleic acid, methyl ester
2074
26.15 ± 0.26
Oleic acid, methyl ester
2099
1.83 ± 0.01
Stearic acid, methyl ester
2212
2.07 ± 0.00
Arachidonic acid methyl ester
2217
6.99 ± 0.01
EPA (methyl ester) – omega-3
2365
9.54 ± 0.12
DHA (methyl ester) – omega-3
16.27
SFA
40.11
MUFA
24.18
PUFA (~16.5% omega-3)
GC-MS analysis of Danube asp oil (derivatized to methyl esters)

16. Methods, Results and Discussion:
GC-MS analysis & Thermal analysis (TG-DTG & DSC)
(Asp oil and its β-cyclodextrin complex)
See: Poster No 14
TG-DTG (top) and DSC (right) analyses of β-cyclodextrin / Danube asp oil complexes at 1:1 molar ratio

17. Principal Component Analysis (PCA)
Scores plots (top) and Loadings plot (right) from the PCA analysis of thermal analyses (TG and DSC) data for β-cyclodextrin/natural highly hydrophobic mixtures (F – fruit oil,
V – vegetable oil, DF – Danube fish oil)

18. Conclusion
TG-DTG and DSC analyses of highly hydrophobic natural compounds mixture (oils)/cyclodextrin complexes reveals three important regions:
(1) up to ~140 °C, where the crystallization water molecules are released,
(2) ~140 to ~275 °C, where other encapsulated molecules can be released,
(3) >275 °C, where the degradation of cyclodextrin / guest compounds are degraded.
the “hydration” water is especially “surface” water after nanoencapsulation of fatty acid glyceride-containing vegetable and fruit oils, while high content of long chain omega-3 fatty acid glycerides (e.g., EPA and DHA) provide complexes with important ratio of “strongly retained” water in the cyclodextrin/oil complex.

19. Conclusion
DSC reveals an extra endothermic-exothermic peak in comparison with TG, at ~ °C for β-CD, which is related to the crystalline-amorphous transformation of cyclodextrin. The disappearance of this peak indicates that all cyclodextrin molecules are involved in the host-guest molecular interaction.
Statistical multivariate analysis technique (PCA) indicates a relationship between TG-DTG and DSC “hydration” water related parameters and the overall composition of the cyclodextrin/oil complex, regarding the saturated, monounsaturated and polyunsaturated (including omega-3) fatty acid glycerides.
More experiments in terms of using standard fatty acid glycerides in the cyclodextrin encapsulation process and other analysis techniques (e.g., FT-IR, XRD, NMR) are needed to support the thermal analysis based observations for these cyclodextrin/highly hydrophobic compound mixtures complexes.

20. Thanks to:
Prof. Dr. Sophie FOURMENTIN (Université du Littoral-Côte d'Opale, Dunkerque, France)
Prof. Dr. Virgil PĂUNESCU (Regional Centre for Immunology and Transplant &
OncoGen Centre – Timişoara)
Dr. Alexandra LUKINICH-GRUIA (Regional Centre for Immunology and Transplant &

21. THANK YOU!