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THE PHYSICAL PROPERTIES OF TRIACYLGLYCEROLS IN RELATION TO HILKFAT
A thesis presented for the degree of Doc t or of Philosophy
in Chemistry at Massey University
by
Robert Norris 1977
ABSTRACT
Enantiome ric and racemic triacylglyce rols (TGs) rep re senta tive of the major structural classe s in milkfat we re synthe sise d and the ir polymorphism was characte rise d by diffe rential scanning calorime try (DSC ) and infrare d (IR) spe ctroscop y.
With butyric (B) , ole ic (0), palmitic (P) and stearic (S) a cids
as starting mate rials, 22 racemic TGs we re prepare d. The main TG classe s we re : 1) palmitoyl-stearoyl TGs (e.g. PSS ), 2) 1-butyryl TGs (BPS) , 3) 1-ole oyl TGs (OPS) , 4) 2-ole oyl TGs ( POS ), 5) 1-butyryl-2-ole oyl TGs (BOS) and 6) 1, 2-diole oyl TGs (OOS) . Three TGs contain ing e la idic acid (E) we re also synthe sise d (BES, ESS and SES). In a ddition, enan
tiome rs of three of the racem ic TGs be lon ging to the 1-butyryl and 1-ole oyl classe s we re prepare d (�-SSB, -SSO an d -PPO).
The polymorphic forms of each TG we re classifie d as a, �· or�
by comparison of the ir IR spe ctra with the spe ctra of the polymorphic forms of monoacid TGs. Solvent crystallise d forms we re also chara cte rise d by X-ray powde r diffraction. Ne lting poin ts of all polymorphs and heats of fusion of the least stable (a) and most stable forms we re de te rmine d by DSC . Howe ve r, the de te rmination of the p olym orphic a ssignment and heat of fusion of the inte rme diate forms wa s often un ce rtain be cause of
the difficulty in obtaining a pure phase.
The principal findings we re :-
1) C orre spondin g enantiome ric and racemic TGs e xhibite d similar p oly
m orphic be haviour e xcept tha t the a forms of the enan tiome rs trans
forme d more rapidly than th ose of the ir ra cemate s.
2) For TGs in which one fatty a cid was ve ry diffe rent from the othe r two (e . g. BSS, OOS) , the position of the un usual acid de te rmine d the chain pack ing of the stable form . If the a cid was in a primary position,
the TG wa s �'-stable (e.g. BSS, OSS, OOS) , while if it was in the se condary position , the TG wa s �-stable (e . g. SOS, SBS).
3 ) The re we re close paralle ls be tween the stable forms of corre sponding
butyryl and ole oyl TGs (e . g. BSS, OSS; SBS, SOS; BOS, OOS), although in othe r re spe cts the ir p olymorph ism ha d little in common . The stable form s of BSP and OSP showe d anomalous the rmal, diffra ction and spe ctral data compare d with the rema ining 1-butyryl and 1-ole oyl TGs.
4 ) The re sults obtaine d for the 1-ole oyl, 2-ole oyl and 1, 2-diole oyl TGs
we re in gene ral a greement with earlie r reports, a lthough some
diffe rences we re note d in the transforma tion of OSP, OPS, SOS and POP.
Furthermore, previously undetected tran sitions were observed for a ll the oleoyl TGs, a lthough these were minor. A new polymorph of OPP was a lso characterised. With the exception of POS, all monooleoyl TGs showed an omalous crysta llisation beha viour.
iii
5) The results for the polymorp hism of the palmitoyl-stearoyl and elaidoyl
stearoyl TGs were a lso in a ccord with p revious reports. The presence of a � ·2 form was confirmed for all TGs except SPS and PSP. The heats of fusion of the � forms of SPS an d PSP were comparable with tho se of their unsymmetrica l counterparts, PSS an d PPS, but the heats of fusion of their stable�· forms were much higher than those of� · SSS, PSS, PPS and PPP.
These findings are discussed in rela tion to the prin ciples of close packin g and their relevan ce to the pha se behaviour of milk fat.
AC KNOWLEDGE1ENTS
I would particularly like to thank my two supe rvisors, Dr J.C . Hawke and Prof. G. N. Malcolm, for the opportunity to work at Masse y Unive rsity. I am also inde bte d to the Ne w Zealand Dairy Re search Institute for financial support during the course of this work.
In the preparation of the the sis, thanks are due to Mrs . A.
Stre tton for drawing m ost of the figure s, to M iss V. L . Be the ll and Dr D . A. D . Parry for proof-reading and to Mrs J. R. Parry for typ ing.
I am similarly obligate d to Me ssrs P. T. Tuttiet t and D. S. M unro, who wrote the compute r p rograms for graphical pre sentation of the the rmal analysis data. Dr M.W. Taylor made helpful sugge stion s of a gene ral nature .
Finally, I am grate ful to m y wife an d family for the ir con tin ue d inte re st and encouragement ove r what must have seeme d an in te rminable pe riod.
C hapte r 1.
1.1.
1. 1. 1 1. 1. 2
1. 1. 3 1. 1. 4 1. 1. 5 1.1. 6
1. 2
1. 2. 1 1. 2. 2 1. 3 1. 3. 1
1. 4 1. 4. 1
C hap te r 2.
A
2. 1 2. 1. 1 2. 1. 2 2. 1. 3 2. 1. 4
2. 2 2. 2. 1 2.2. 2 2. 2. 3 2. 2. 4
TABLE OF CONTENTS
INTRODUCTION
Polymorphism of Triacylglyce rols Historical Backgroun d
Structural Fe ature s of Lipids in the Solid State
Monoacid Saturate d Triacylglyce rols Unsaturate d Triacylglyce rols
Diacid Saturate d Triacylglyce rols Enan tiome ric Triacylglyce rols
Infrare d Spe ctroscop y of Triacylglyce rols in the Solid State
Assignments of C haracte ristic Absorp tion Bands Spe ctral Diffe rence s be tween Polym orphic Forms
Me lting and Solidification of Fats
Te chnical Imp ortance of the Phase Behaviour of Fats
Polym orphism of Fats Solid Miscibility of Fats
He lting and Solidification of Hilkfat and Milkfat Fraction s
Aim of the Pre sent Work
Se le ction of Triacylglyce rols for Synthe sis HATERIALS AND HE.rHODS
Analytical and Synthe tic Me thods Mate rials
Solvents Re age nts Fatty Acids
Storage of Glyce ride s and Syn the tic In te rme diate s
Analytical Me thods
Thin-Laye r C hrom atography Column Chromatography Gas-Liquid C hromatography
Structural Analysis of Triacylglyce rols
V
1
1 1 2
5 13 18 21
22
22 24
27 27
27 28 29
33 33
36 36 36 36 36 37 37
37 37 39 40 40
2. 3 2. 3. 1 2. 3. 2 2. 3. 3 2. 3. 4 2. 3. 5
2. 4 2. 4. 1 2. 4. 2 2. 4. 3
B
2. 5 2. 6 2. 6. 1 2. 6. 2 2. 6. 3 2. 7 2. 7. 1 2. 7. 2 2. 7. 3
2. 8 2. 8. 1 2. 8. 2 2. 8. 3 2. 8. 4 2. 8. 5
2. 9
C hapter 3.
3. 1 3. 2 3. 2. 1
Preparation of Racemic Triacylglycerols Acyl Chlorides
1-Acylglycerols 1,3-Diacylglycerols 1,2-Distearoylglycerol Triacylglycerols
Preparation of Enantiomeric Triacylglycerols 1,2-Isopropylidene-�-glycerol
1,2-Diacyl-�-glycerols Triacylglycerols
Physical M ethods
M elting Point Determination X-Ray Powder Diffraction
Apparatus
Preparation of Samples
Identification of Polym orphs
In frared Spectroscopy Apparatus
Investigation of Polymorphism Identification of Polymorphs Thermal Analysis
Apparatus C alibration
Presentation of Data
Investigation of Polymorphism Identification of Polymorphs Preparation of� PSP and�· SPS
RESULTS
Structural Analysis of Triacylglycerols Polymorphism of Racemic Triacylglycerols Palmitoyl-Stearoyl and Elaidoyl-Stearoyl Triacylglycerols
1-Butyryl-2-0leoyl and 1,2-Dioleoyl Triacylglycerols
1,2-Dibutyryl-3-palmitoylglycerol
40 42 42 44 47 49
50 50 52 53
54 54 54 54 54 54 55 55 55 56
56 56 57 57 59 61
61
62 62 62 62
64
66
C hapte r
3.2.4 3.2.5 3.2.6 3.2.7 3.2.8
4.
4.1 4.1.1
4.1.2
4.1.3 4.1.4 4.1.5 4.1.6 4.1.7 4.1.8 4.2 4.2.1
4.3
2-0le oyl Triacylglyce rols
2-Butyryl-1,3-distearoylglyce rol 1-Butyryl Triacylglyce rols
1-Butyryl-2-e laidoyl-3-ste aroylglyce rol 1-0le oyl Triacylglyce rols
Polymorphism of Enantiome ric Triacylglyce rols Comparison of the Polymorphism of Corre sponding Enantiome ric an d Race mic Triacylglyce rols
DISC USSION
Polymorphism of Race mic Triacylglyce rols Palmitoyl-Ste aroyl and Elaidoyl-Ste aroyl
Triacylglyce rols
1-Butyryl-2-0le oyl and 1,2-Diole oyl Triacylglyce rols
1,2-Dibutyryl-3-palmitoylglyce rol 2-0le oyl Triacylglyce rols
2-Butyryl-1,3-distearoylglyce rol 1-Butyryl Triacylglyce rols
1-Butyryl-2-e laidoyl-3-ste aroylglyce rol 1-0le oyl Triacylglyce rols
Polymorphism of Enantiome ric Triacylglyce rols Comp arison of the Polymorphism of Corre sponding Enan tiome ric and Race mic Triacylglyce rols
Con clusion
APPENDIX
The rmal and Spe ctral Data for the Polymorphic Forms of the Individual Triacylglyce rols
BIBLIOGRAPHY
vii
67 72 73 75 76
81 81
120
120 120
121
123 124 127 128 132 133
136 136
138 140
19 3
Table
1.1 1.2
1.4
1.6
3.4
3-5
3.6
3-7
3.8
3-9
3.10
3.11 3.12
1 2
3
LIST OF TABLES
C lassification of the Polym orphic Forms of Glyce ride s The Polymorphs of 1-0le oyl and 2-0le oyl Disaturate d Triacylglyce rols
C haracte ristic Bands in the Solid State Spe ctra of Triacylglyce rols
8 15
23
M e thylene Waggin g Band Distributions for the Alpha Forms 25 of the Monoacid Triacylglyce rols of Elaidic, Ole ic,
Palmitic and Ste aric Acids
C omparison of the Spe ctra of a, � · and� Polymorphs of 26 M on oacid Saturate d Triacylglyce rols
Selection of Triacylglyce rols for Syn the sis 34
Positional Analysis of Synthe tic Triacylglyce rols 83 Stere ospe cific Analysis of �-SSO and �-SSB 83 X-Ray Short Spacin gs of the Solvent C rystallise d Form s of 84 Palm itoyl-Ste aroyl and Elaidoyl-Ste aroyl Triacylglyce rols
M e lting Points and He ats of Fusion of the Polymorphic 85 Form s of Palmitoyl-Ste aroyl and Elaidoyl-Ste aroyl
Triacylglyce rols
Me lting Points and He ats of Fusion of the Polymorphic 86 Form s of OOS, OOP, BOS, BOP and BBP.
X-Ray Short Sp acings of the Solvent C rystallise d Forms 87 of SOS, POS, POP and SBS
Me lting Poin ts and He ats of Fusion of the Polymorphic 88 Form s of SOS, POS, POP and SBS
X-Ray Short Spacings of the Solvent C rystallise d Forms of 89 BSS, BSP, BPS, BPP and BES
M e lting Points and He ats of Fusion of the Polymorphic 89 Forms of BSS, BSP, BPS 1 BPP and BES
X-Ray Short Spacings of the Solvent C rystallise d Forms 90 of OSS, OSP, OPS and OPP
Melting Points and He ats of Fusion of the Polym orphic 91 Form s of OSS, OSP, OPS and OPP
X-Ray Short Spacings of the Solvent C rystallise d Forms 92 of C orre spondin g Racem ic an d Enantiome ric Triacylglyce rols M e lting Points and He ats of Fusion of the Polym orphic 92
Form s of C orre spondin g Racemic and Enantiome ric Triacylglyce rols
APPENDIX
M e lting Points of the Stable Forms of Triacylglyce role Some Common Bands in the Infrare d Spe ctra of Re late d Stable Forms
Some Common Bands in the Infrare d Spe ctra of Re late d [3 and � 1 Form s
141 142
143
ix
LIST OF FIGURES Figure
1-1 Triclinic parallel and orthorhombic p erp endicular 3 subcells
1-2 The main polym orphic forms of monoacid triacylglycerols 6 1-3 Melting points of monoacid triacylglycerols 9 1-4 Molecular arrangement of trilaurin, b-axis projection 10 1-5 Triacylglycerols in the liquid state, p roposed lam ellar 12
structure
1-6 Trip le chain structure of the � stable form of 2-oleoyl- 16 distearoylglycerol
1�7 Layer stacking of methyl end groups in the � forms of 20 C /C 2 triacylglycerols showing the two terrace structures
n n±
1-8 Triacylglycerols of milkfat 31
2-1 Scheme for the p reparation of oleic acid 38 2-2
2-3 2-4 2-5 2-6
3-1
3-2
3-3 3-4 3-5 3-6 3-7
Gen eral schem e for the synthesis of racemic triacylglycerols 41
Synthesis of racemic triacylglycerols 43
Synthesis of racemic 1,2-diacylglycerol 48 Synthesis of enantiomeric triacylglycerols 51 C on ventions for the presentation of thermal analysis data 58 X-Ray diffraction patterns of the � stable forms of
p almitoyl-stearoyl and elaidoyl-stearoyl triacylglycerols X-Ray diffraction p atterns of the� and�· forms of PSP and SPS
Melting therm ograms of � SSS, SPS, PPP and SES Melting thermograms of � PSP, PSS, PPS and ESS Melting therm ograms of �· sss, ppp and SES Melting thermograms of �I PSS, PPS and ESS
Melting therm ograms of the solven t crystallised �I and
� forms of SPS and PSP
93
94
9 5 9 6 9 7 9 8 9 9
3-8 IR spectra of the �1 stable forms of OOS, OOP, BOS and BOP 100 3-9 Melting therm ograms of � OOS, OOP, BOS and BOP 101 3-10 X-Ray diffraction p atterns of the � stable forms of SOS, 102
POS , POP and SBS
3-11 IR spectra of the � stable forms of SOS, POS, POP, SBS, 103 SES and ESS
3-12 IR spectra of comparative �2 and�� p olymorp hs of SOS, 104 POS and POP
3-13 3-14
3-15
Melting therm ograms of � SOS, POS and POP
Melting therm ograms of the intermediate form s of SOS, POS and POP
Polymorphism of SOS, POS and POP
105 106
107
Figure 3-16
3-17
3-18 3-19
3-20
3-21
3-22
3-23
3-24 3-25
3-26
3-27
4-1
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17
Page IR spectra of the�· stable forms of OXY and BXY 108 triacylglycerols
X-Ray diffraction patterns of the�' stable forms of OXY 109 and BXY triacylglycerols
Melting thermograms of a BSS, BSP, BPS and BPP 110 Melting thermograms of the intermediate forms of BSS, BSP, 111 BPS and BPP
Melting thermograms of a OSS, OSP, OPS and OPP showing 112 the presence of the a2 form at high heating rates
Melting thermograms of a OSS, OSP, OPS and OPP recorded 113 after tempering at the a2 peak temperature
Melting thermograms of a OSS, OSP, OPS and OPP recorded 114 at low heating rates
Melting thermograms of the intermediate forms of OSS, OSP, 115 OPS and OPP
Polymorphism of OSS, OSP, OPS and OPP 116 IR spectra of the�· stable forms of racemic and 117 enantiomeric triacylglycerols
X-Ray diffraction patterns of the stable forms of corres- 118 ponding racemic and enantiomeric triacylglycerols
Helting thermograms of the a forms of rac- and sn-SSB and 119 SSO showing the more rapid transformatiOn of the-antipodes
Comparison of glycerol conformations in "tuning-fork" and 130
"chair" structures
Thermal behaviour Thermal behaviour Thermal behaviour Thermal behaviour Thermal behaviour Thermal behaviour Thermal behaviour IR spectra of SES Thermal behaviour IR spectra of ESS Thermal behaviour IR spectra of OOS Thermal behaviour IR spectra of OOP Thermal behaviour IR spectra of BOS Thermal behavi·our
of sss of PSS of PPS of ppp of SPS of PSP of SES of ESS of OOS of OOP of BOS of BOP
APPENDIX
144 145 146 147 148 149 150 151 152 153 154 155 156 157 158 159 160
xi
Figure Page
18 IR spectra of BOP 161
19 Thermal behaviour of BBP 162
20 IR spectra of BBP 163
21 Thermal behaviour of SOS 164
22 Crystallisation of SOS from the melt 165
23 IR spectra of SOS 166
24 Thermal behaviour of POS 167
25 IR spectra. of POS 168
26 Thermal behaviour of POP 169
27 IR spectra of POP 170
28 Thermal behaviour of SBS 171
29 IR spectra of SBS 172
30 Thermal behaviour of sn-SSB 173
31 Thermal behaviour of BSS 174
32 IR spectra of BSS 175
33 Thermal behaviour of BSP 176
34 IR spectra of BSP 177
35 Thermal behaviour of BPS 178
36 Thermal behaviour of BPP 179
37 IR spectra of BPP 180
38 Thermal behaviour of BES 181
39 IR spectra of BES 182
40 Thermal behaviour of sn-SSO 183
41 Thermal behaviour of OSS 184
42 IR spectra of OSS 185
43 Thermal behaviour of OSP 186
44 IR spectra of OSP 187
45 Thermal behaviour of OPS 188
46 IR spectra of OPS 189
47 Thermal behaviour of sn-PPO 190
48 Thermal behaviour of OPP 191
49 IR spectra of OPP 192
NOMENCLATURE
