NZ743612A - Method for selectively synthesizing cationic lipids - Google Patents
Method for selectively synthesizing cationic lipidsInfo
- Publication number
- NZ743612A NZ743612A NZ743612A NZ74361216A NZ743612A NZ 743612 A NZ743612 A NZ 743612A NZ 743612 A NZ743612 A NZ 743612A NZ 74361216 A NZ74361216 A NZ 74361216A NZ 743612 A NZ743612 A NZ 743612A
- Authority
- NZ
- New Zealand
- Prior art keywords
- cationic lipid
- fatty acid
- formula
- preparing
- carbon atoms
- Prior art date
Links
- 125000002091 cationic group Chemical group 0.000 title claims abstract description 15
- 230000002194 synthesizing Effects 0.000 title abstract description 17
- -1 cationic lipid Chemical class 0.000 claims abstract description 59
- 125000004432 carbon atoms Chemical group C* 0.000 claims abstract description 32
- 229910052739 hydrogen Inorganic materials 0.000 claims abstract description 21
- 239000001257 hydrogen Substances 0.000 claims abstract description 21
- 125000005313 fatty acid group Chemical group 0.000 claims abstract description 19
- 125000005314 unsaturated fatty acid group Chemical group 0.000 claims abstract description 16
- 125000005471 saturated fatty acid group Chemical group 0.000 claims abstract description 13
- 125000004435 hydrogen atoms Chemical group [H]* 0.000 claims abstract description 9
- 238000006243 chemical reaction Methods 0.000 claims abstract description 7
- 235000014113 dietary fatty acids Nutrition 0.000 claims description 21
- 239000000194 fatty acid Substances 0.000 claims description 21
- 150000002632 lipids Chemical class 0.000 claims description 20
- VILCJCGEZXAXTO-UHFFFAOYSA-N Triethylenetetramine Chemical compound NCCNCCNCCN VILCJCGEZXAXTO-UHFFFAOYSA-N 0.000 claims description 12
- 125000002811 oleoyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])/C([H])=C([H])\C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 11
- 239000003960 organic solvent Substances 0.000 claims description 11
- RTZKZFJDLAIYFH-UHFFFAOYSA-N diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 claims description 8
- 150000002430 hydrocarbons Chemical class 0.000 claims description 7
- UFHFLCQGNIYNRP-UHFFFAOYSA-N hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 claims description 7
- 150000002431 hydrogen Chemical group 0.000 claims description 5
- 239000004215 Carbon black (E152) Substances 0.000 claims description 4
- 239000002253 acid Substances 0.000 claims description 4
- 125000003312 cerotoyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 4
- 239000011780 sodium chloride Substances 0.000 claims description 4
- AQGNVWRYTKPRMR-UHFFFAOYSA-N N'-[2-[2-[2-[2-(2-aminoethylamino)ethylamino]ethylamino]ethylamino]ethyl]ethane-1,2-diamine Chemical compound NCCNCCNCCNCCNCCNCCN AQGNVWRYTKPRMR-UHFFFAOYSA-N 0.000 claims description 3
- 238000007792 addition Methods 0.000 claims description 3
- FAGUFWYHJQFNRV-UHFFFAOYSA-N tetraethylenepentamine Chemical compound NCCNCCNCCNCCN FAGUFWYHJQFNRV-UHFFFAOYSA-N 0.000 claims description 3
- RPNUMPOLZDHAAY-UHFFFAOYSA-N DETA Chemical compound NCCNCCN RPNUMPOLZDHAAY-UHFFFAOYSA-N 0.000 claims description 2
- LSHROXHEILXKHM-UHFFFAOYSA-N N'-[2-[2-[2-(2-aminoethylamino)ethylamino]ethylamino]ethyl]ethane-1,2-diamine Chemical compound NCCNCCNCCNCCNCCN LSHROXHEILXKHM-UHFFFAOYSA-N 0.000 claims description 2
- 150000001335 aliphatic alkanes Chemical group 0.000 claims description 2
- 125000000217 alkyl group Chemical group 0.000 claims description 2
- 125000002886 arachidonoyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])/C([H])=C([H])\C([H])([H])/C([H])=C([H])\C([H])([H])/C([H])=C([H])\C([H])([H])/C([H])=C([H])\C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 2
- 125000001124 arachidoyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 2
- 125000003910 behenoyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 2
- 125000001901 erucoyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])/C([H])=C([H])\C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 2
- 125000000400 lauroyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 2
- 125000002669 linoleoyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])/C([H])=C([H])\C([H])([H])/C([H])=C([H])\C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 2
- 239000000463 material Substances 0.000 claims description 2
- 125000000265 myristoleoyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])/C([H])=C([H])\C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 2
- 125000001419 myristoyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 2
- 230000003472 neutralizing Effects 0.000 claims description 2
- 125000001236 palmitoleoyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])/C([H])=C([H])\C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 2
- 125000001312 palmitoyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 2
- 150000003839 salts Chemical class 0.000 claims description 2
- 125000003696 stearoyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 2
- IMENJLNZKOMSMC-UHFFFAOYSA-N N'-[2-[2-[2-[2-[2-(2-aminoethylamino)ethylamino]ethylamino]ethylamino]ethylamino]ethyl]ethane-1,2-diamine Chemical compound NCCNCCNCCNCCNCCNCCNCCN IMENJLNZKOMSMC-UHFFFAOYSA-N 0.000 claims 1
- QYDYPVFESGNLHU-KHPPLWFESA-N methyl oleate Chemical compound CCCCCCCC\C=C/CCCCCCCC(=O)OC QYDYPVFESGNLHU-KHPPLWFESA-N 0.000 description 14
- 230000015572 biosynthetic process Effects 0.000 description 11
- 238000001308 synthesis method Methods 0.000 description 11
- 238000003786 synthesis reaction Methods 0.000 description 11
- VLKZOEOYAKHREP-UHFFFAOYSA-N hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 10
- HEMHJVSKTPXQMS-UHFFFAOYSA-M sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 9
- 238000005160 1H NMR spectroscopy Methods 0.000 description 8
- 239000003814 drug Substances 0.000 description 8
- 229940079593 drugs Drugs 0.000 description 8
- 150000004665 fatty acids Chemical class 0.000 description 7
- 229940073769 methyl oleate Drugs 0.000 description 7
- 239000000203 mixture Substances 0.000 description 7
- 238000005227 gel permeation chromatography Methods 0.000 description 6
- IJGRMHOSHXDMSA-UHFFFAOYSA-N nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 6
- 125000000129 anionic group Chemical group 0.000 description 5
- 239000008079 hexane Substances 0.000 description 5
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 description 4
- 125000003277 amino group Chemical group 0.000 description 4
- 239000007795 chemical reaction product Substances 0.000 description 4
- HEDRZPFGACZZDS-UHFFFAOYSA-N chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 4
- 238000000746 purification Methods 0.000 description 4
- FAPWRFPIFSIZLT-UHFFFAOYSA-M sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- 238000003756 stirring Methods 0.000 description 4
- 125000005907 alkyl ester group Chemical group 0.000 description 3
- 150000001768 cations Chemical class 0.000 description 3
- BDAGIHXWWSANSR-UHFFFAOYSA-N formic acid Chemical compound OC=O BDAGIHXWWSANSR-UHFFFAOYSA-N 0.000 description 3
- 150000002500 ions Chemical class 0.000 description 3
- 238000000034 method Methods 0.000 description 3
- 229910052757 nitrogen Inorganic materials 0.000 description 3
- 150000007523 nucleic acids Chemical class 0.000 description 3
- 108020004707 nucleic acids Proteins 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 2
- HEDRZPFGACZZDS-MICDWDOJSA-N cdcl3 Chemical compound [2H]C(Cl)(Cl)Cl HEDRZPFGACZZDS-MICDWDOJSA-N 0.000 description 2
- 238000005119 centrifugation Methods 0.000 description 2
- 239000000839 emulsion Substances 0.000 description 2
- 230000002209 hydrophobic Effects 0.000 description 2
- 230000003993 interaction Effects 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 238000004949 mass spectrometry Methods 0.000 description 2
- OKKJLVBELUTLKV-UHFFFAOYSA-N methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 2
- YMWUJEATGCHHMB-UHFFFAOYSA-N methylene dichloride Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 2
- 239000000693 micelle Substances 0.000 description 2
- 239000002105 nanoparticle Substances 0.000 description 2
- 150000003141 primary amines Chemical class 0.000 description 2
- 150000003335 secondary amines Chemical group 0.000 description 2
- 210000001124 Body Fluids Anatomy 0.000 description 1
- 210000000170 Cell Membrane Anatomy 0.000 description 1
- VEXZGXHMUGYJMC-UHFFFAOYSA-N HCl Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 1
- 241000282619 Hylobates lar Species 0.000 description 1
- 241000229754 Iva xanthiifolia Species 0.000 description 1
- RUSNFULRUJHOPI-UHFFFAOYSA-N N'-[2-[2-[2-[2-[2-[2-(2-aminoethylamino)ethylamino]ethylamino]ethylamino]ethylamino]ethylamino]ethyl]ethane-1,2-diamine Chemical compound NCCNCCNCCNCCNCCNCCNCCNCCN RUSNFULRUJHOPI-UHFFFAOYSA-N 0.000 description 1
- JXDNGTCNFHAVIO-UHFFFAOYSA-N N,N-dimethylmethanamine;oxolane Chemical compound CN(C)C.C1CCOC1 JXDNGTCNFHAVIO-UHFFFAOYSA-N 0.000 description 1
- 238000005481 NMR spectroscopy Methods 0.000 description 1
- 230000002378 acidificating Effects 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 150000001412 amines Chemical class 0.000 description 1
- QGZKDVFQNNGYKY-UHFFFAOYSA-O ammonium Chemical compound [NH4+] QGZKDVFQNNGYKY-UHFFFAOYSA-O 0.000 description 1
- 239000010839 body fluid Substances 0.000 description 1
- 210000004027 cells Anatomy 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 230000003013 cytotoxicity Effects 0.000 description 1
- 231100000135 cytotoxicity Toxicity 0.000 description 1
- 238000010494 dissociation reaction Methods 0.000 description 1
- 230000005593 dissociations Effects 0.000 description 1
- 238000004821 distillation Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000002708 enhancing Effects 0.000 description 1
- 235000019253 formic acid Nutrition 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- 229910000041 hydrogen chloride Inorganic materials 0.000 description 1
- 230000003834 intracellular Effects 0.000 description 1
- 125000000403 lignoceroyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 1
- 239000002502 liposome Substances 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- IMNFDUFMRHMDMM-UHFFFAOYSA-N n-heptane Chemical compound CCCCCCC IMNFDUFMRHMDMM-UHFFFAOYSA-N 0.000 description 1
- 230000001264 neutralization Effects 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- 230000001376 precipitating Effects 0.000 description 1
- 239000011541 reaction mixture Substances 0.000 description 1
- 238000010992 reflux Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 238000004611 spectroscopical analysis Methods 0.000 description 1
- 125000001424 substituent group Chemical group 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
Abstract
Disclosed are methods for preparing a cationic lipid represented by Formula 1, wherein in Formula 1, a and b are independently 1 to 6, and n and m are independently 0 to 12, with the proviso that 1 ≤ n + m ≤ 12; and wherein one of the following (a) to (c) applies: (a) one of R1 and R4 is a saturated or unsaturated fatty acid group having 12 to 26 carbon atoms, and the other of R1 and R4 is hydrogen, and R2 and R3 are each independently hydrogen; or (b) R1 and R4 are each independently a saturated or unsaturated fatty acid group having 12 to 26 carbon atoms, and R2 and R3 are each independently hydrogen, or (c) R1, R2, R3 and R4 are each independently a saturated or unsaturated fatty acid group having 12 to 26 carbon atoms. The disclosed methods are capable of selectively synthesizing cationic lipids of Formula 1 by controlling the introduction rate of a fatty acid group with respect to an oligoalkyleneamine by the change of reaction conditions. ed or unsaturated fatty acid group having 12 to 26 carbon atoms, and the other of R1 and R4 is hydrogen, and R2 and R3 are each independently hydrogen; or (b) R1 and R4 are each independently a saturated or unsaturated fatty acid group having 12 to 26 carbon atoms, and R2 and R3 are each independently hydrogen, or (c) R1, R2, R3 and R4 are each independently a saturated or unsaturated fatty acid group having 12 to 26 carbon atoms. The disclosed methods are capable of selectively synthesizing cationic lipids of Formula 1 by controlling the introduction rate of a fatty acid group with respect to an oligoalkyleneamine by the change of reaction conditions.
Description
VERIFICATION 0F ATION
PCT International ation No.
I, Junghee LEE (name of translator)
Of 115 Teheran—ro, Gangnam—gu, Seoul 06134, Republic of Korea ss of translator)
hereby declare as follows:
1. I am well acquainted with both the English and Korean languages, and
2. I am the translator of the documents attached hereto and certify that the following is a
true and correct translation of PCT Application No. PCT/ICRZOIG/OISSZS filed
on December 29, 2016, to the best ofmy knowledge and belief.
Dated this l¢7+ day of 7am.2018
S—%‘ignature 0ftranslator
【DESCRIPTION】
【TITLE OF THE INVENTION】
METHOD FOR SELECTIVELY SYNTHESIZING CATIONIC LIPIDS
【Technical Field】
The present disclosure relates to a synthesis method that allows control of the
introduction rate and introduction position of a fatty acid group which is introduced into an
oligoalkyleneamine during synthesis of cationic lipids.
【Background Art】
Until now, the synthesis of cationic lipids in which saturated or unsaturated fatty acid
groups are uced into an amine group of the oligoalkyleneamine has been reported to
introduce lipids into primary amines at both ends of the oligoalkyleneamine (see US Patent
No. 9,220,779, US Patent No. 5,744,355, etc.). However, under the synthesis ions of the
prior art, since the fatty acid groups react nonspecifically with primary and secondary amine
groups of the oligoalkyleneamine, it is ible to react lipids selectively with only amine
groups at one or both ends of the oligoalkyleneamine by such conventional synthesis methods.
Therefore, a mixture having ent lipid introduction rates is synthesized, and a mixture
having ent cationic lipid compositions can be synthesized for each on. It is very
difficult to separate and purify the thus synthesized mixture of cationic lipids into lipids
having the same introduction rates, respectively, and there is a problem that many processes
are required. Therefore, there is a need for a method that can e an oligoalkyleneaminebased
cationic lipid in an environmentally-friendly and ical manner, and can
selectively introduce lipids into an amine group.
【DETAILED DESCRIPTION OF THE INVENTION】
【Technical Problem】
Under these circumstances, the present inventors have conducted intensive studies on
selective synthesis methods capable of introducing fatty acid groups into oligoalkyleneamines
at desired positions and introduction rates during the synthesis of cationic lipids as described
above. As a result, the inventors have unexpectedly found that, when changing conditions of
oligoalkyleneamine and fatty acid tive to be reacted, it is possible to obtain a cationic
lipid having the desired introduction rate and position of fatty acid groups in a simple,
economical and environmentally friendly manner, y completing the present invention.
In view of the above, one object of the present invention is to provide a synthesis
method of a cationic lipid ented by Formula 1 that can selectively introduce a fatty acid
group into a y or secondary amine group of oligoalkylene amine and can control the
introduction rate of the fatty acid to be introduced.
Another object of the present invention is to provide a method capable of efficiently
purifying cationic .
[Formula 1]
in the above formula, the definition of the substituents is as defined below.
【ADVANTAGEOUS EFFECTS】
The method for synthesizing a cationic lipid according to the present invention can
control the introduction rate of the fatty acid group to the oligoalkyleneamine by merely
adjusting the synthesis conditions unlike a conventional . Therefore, unlike the
conventional method in which a mixture of ic lipids having ent lipid introduction
rates are synthesized at the time of synthesis, since it is possible to synthesize a cationic lipid
which consistently has high purity and uniform introduction rate, purification process with
high difficulty is unnecessary. In addition, the synthesis and cation steps are simple, and
economical efficiency in mass production is high. Thus, it is very useful for forming an
intracellular delivery complex capable of enhancing stability in body fluids together with
anionic drugs such as nucleic acid or anionic active ingredients, or for preparing cationic
lipids capable of forming liposomes, micelles, emulsions, and nanoparticle drug delivery
system.
【BRIEF DESCRIPTION OF GS】
shows the s of measurement (GPC) of the change in the molecular
weight according to the change in the number of fatty acid groups.
shows the results of proton nuclear magnetic resonance spectroscopy (1H
NMR) analysis of 1,6-dioleoyl triethylenetetramide.
shows the results of proton nuclear magnetic resonance spectroscopy (1H
NMR) is of tetraoleoyl triethylenetetramide.
【DETAILED DESCRIPTION OF THE EMBODIMENTS】
In one aspect for achieving the above object, the present invention relates to a method
capable of synthesizing a cationic lipid represented by Formula 1 with high purity by
controlling the uction rate of a fatty acid group.
[Formula 1]
Specifically, the present invention is characterized by reacting an oligoalkyleneamine
ented by a 2 with a fatty acid alkyl ester represented by a 3.
[Formula 2]
[Formula 3]
in the above ae 1 to 3,
n and m are independently 0 to 12, with the proviso that 1 ≤ n + m ≤ 12,
a and b are independently 1 to 6,
R1, R2, R3 and R4 are independently hydrogen or saturated or unsaturated fatty acid
group having 12 to 26 carbon atoms, with the proviso that at least one of R1 and R4 is
saturated or unsaturated fatty acid group having 12 to 26 carbon atoms,
R is saturated or unsaturated hydrocarbon having 11 to 25 carbon atoms, and
R5 is an alkyl group having from 1 to 14 carbon atoms.
In order to keep the density of the fatty acid group high and to ze the
cytotoxicity induced by cations, it is preferable that n and m have the numerical value and
range as described above.
In addition, with respect to the R and R1 to R4, if the number of carbon atoms in the
saturated or unsaturated hydrocarbon is less than 11, the hydrophobic interaction between the
hydrocarbon chains can decrease, and thus a ation stable with the anionic drug cannot
be formed. On the other hand, if the number of carbon atoms is larger than 25, the
hydrophobic interaction between the hydrocarbons will increase, and thus a formulation
excessively stable with the c drug will form, whereby the in vivo dissociation of the
drug will decrease, leading to a decrease in the efficacy of the drug. In addition, the ure
of the hydrocarbon chains will increase due to an increase in cis double bonds, and thus the
resulting ation will have low density and thus low stability.
In a preferred embodiment, in the ive synthesis method according to the present
invention, a cationic lipid of Formula 1 wherein one of R1 and R4 is hydrogen and R2 and R3
are each hydrogen can be prepared by ing the molar ratio (oligoalkyleneamine/fatty acid
alkyl ester) of the oligoalkyleneamine of Formula 2 to fatty acid alkyl ester of Formula 3 to
more than 1 to 20 or less, preferably 3 or more to 8 or less.
In another preferred embodiment, in the selective synthesis method according to the
present invention, a cationic lipid of Formula 1 n R1 and R4 are fatty acid groups
having 12 to 26 carbon atoms and R2 and R3 are hydrogen can be prepared by adjusting the
molar ratio of the fatty acid alkyl ester of Formula 3 to the oligoalkyleneamine of Formula 2
to 1 or more to 5 or less, preferably 1.5 or more to 4 or less.
In r preferred embodiment, in the selective synthesis method according to the
present invention, a cationic lipid of Formula 1 wherein R1, R2, R3 and R4 are a fatty acid
group having 12 to 26 carbon atoms can be prepared by adjusting the molar ratio of the fatty
acid alkyl ester of Formula 3 to the lkyleneamine of Formula 2 to more than 5 to 20 or
less, preferably 6 or more to 10 or less.
In the above-described synthesis method according to the present invention, the
reaction is carried out without using an organic solvent during the reaction of the
oligoalkyleneamine with the fatty acid alkyl ester.
In yet another , the present invention provides a method for preparing a cationic
lipid of Formula 1 wherein R1 and R4 are a fatty acid group having 12 to 26 carbon atoms
and one of R2 and R3 is hydrogen, the method comprising a step of reacting the cationic lipid
of Formula 1 n R1 and R4 are a fatty acid group having 12 to 26 carbon atoms and R2
and R3 are en with a fatty acid alkyl of Formula 3 to prepare a cationic lipid of
Formula 1 wherein R1 and R4 are a fatty acid group having 12 to 26 carbon atoms and one of
R2 and R3 is en.
Preferably, n and m are independently 0 to 9, with the proviso that 1 ≤ n+m ≤ 10.
Preferably, a and b may be 2 to 4.
Preferably, R1, R2, R3, and R4 may be independently unsaturated fatty acid group
having 14 to 22 carbon atoms.
ably, one or more of R1, R2, R3 and R4 may be selected from the group
consisting of lauroyl, myristoyl, palmitoyl, stearoyl, arachidoyl, behenoyl, lignoceroyl,
cerotoyl, myristoleoyl, palmitoleoyl, sapienoyl, oleoyl, linoleoyl, arachidonoyl,
eicosapentaenoyl, erucoyl, docosahexaenoyl, and cerotoyl.
In the process for preparing a cationic lipid of a 1 wherein R1 and R4 are fatty
acid groups having 12 to 26 carbon atoms and one of R2 or R3 is hydrogen, it is desirable
that the molar ratio of the fatty acid alkyl ester to the cationic lipid of Formula 1 wherein R1
and R4 are fatty acids having 12 to 26 carbon atoms and R2 and R3 are hydrogen is 0.5 or
more to 20 or less, preferably 0.7 or more to 10 or less, more preferably 1 or more to 5 or
less.
In the present invention, the oligoalkyleneamine of Formula 2 is specifically
oligoethyleneamine. More specifically, it may be at least one selected from the group
consisting of diethylenetriamine, triethylenetetramine, tetraethylenepentamine,
thylenehexamine, hexaethyleneheptamine, thyleneoctamine,
octaethylenenonamine, nonaethylenedecamine, decaethyleneundecamine,
undecaethylenedodecamine, dodecaethylenetridecamine and tridecaethylenetetradecamine,
but is not d thereto. Preferably, it is at least one selected from the group consisting of
triethylenetetramine, tetraethylenepentamine, pentaethylenehexamine and
hexaethyleneheptamine.
As bed above, when the oligoalkyleneamine of Formula 2 and the fatty acid
alkyl ester of Formula 3 are reacted at the above equivalent ratio, a high-purity ic lipid
can be synthesized by adjusting the hydrocarbon introduction rate in the produced cationic
lipid.
According to the method of the present invention, the cationic lipid can be easily
synthesized at a high yield by using a fatty acid derivative such as inexpensive
oligoalkyleneamine and fatty acid alkyl ester, which is environmentally friendly and
economical. In addition, it is advantageous in that the lipid synthesized through the above
on has a low lity in a nonpolar organic t and thus is easily precipitated, so
that the purification process of the synthesized product is very simple.
Therefore, in another preferred embodiment, the present invention may further
include a step of adding a nonpolar organic solvent to the cationic lipid of Formula 1
produced by the above synthesis method, precipitating and separating unreacted materials to
purify the cationic lipid. Preferably, the nonpolar organic solvent may be an alkane or ether
having 4 to 12 carbon atoms, more preferably , heptane or l ether, but is not
limited thereto.
In another preferred embodiment, the present invention may further include a step of
dissolving the cationic lipid of Formula 1 produced by the above synthesis method by adding
a nonpolar organic solvent, adding an acid thereto to separate the cationic lipid as an acid
addition salt into the aqueous layer from the organic solvent, neutralizing the separated lipid,
and extracting it with a nonpolar c solvent, followed by separation and cation.
Further, the preferred nonpolar organic solvent may be chloroform or dichloromethane, but is
not limited o.
As described above, since the cationic lipid of a 1 produced by the synthesis
method according to the present invention itself exhibits low solubility, easily precipitates and
exhibits a m introduction rate, the purification method of the present invention using
this point has the advantage in that it is economical, environmentally friendly and simple as
compared with the tional purification method of cationic lipids.
Since the ic lipid synthesized and/or purified according to the present invention
retains a positively charged state in cells because the amine group of the oligoalkyleneamine
exists in a vely charged form at a en ion concentration (pH) of a l region
which is a normal in vivo environment. Therefore, the cationic lipid not only makes it
possible to form a complex with an anionic drug containing a negatively charged nucleic acid
at neutral pH, such as in vivo, and to increase contact with negatively charged target cell
membranes. Thus, the cationic lipids of the present invention can be used to produce various
forms of anionic drug delivery formulations, such as mes, micelles, emulsions, and
nanoparticles for nucleic acid delivery ations.
Hereinafter, the present invention will be described in more detail with reference to
the following examples. However, these examples are provided herein for illustrative
purposes only and should not be used to limit the scope of the present invention in any manner.
Example 1: Synthesis of leoyl triethylenetetramide
.00 g (33.34 mmol) of triethylenetetramine and 2.00 g (6.69 mmol) of methyl oleate
were placed in a round bottom flask and then allowed to react with stirring with a magnetic
bar at 65°C under en for 5 days.
After completion of the on, the reaction product was ved in 150 mL of
diethyl ether, and then sodium chloride (NaCl) was added to 30 mL of 1 M sodium hydroxide
(NaOH) solution in a separating funnel, and the reaction mixture was washed three times to
remove unreacted triethylenetetramine. The upper organic solvent layer in the separating
funnel was heated and distilled under reduced pressure with a distillation condenser.
The finally obtained product was ed by HP1100 series gel tography
using Shodex KF-801 and KF-802 columns in 0.5% v/v trimethylamine-tetrahydrofuran
mobile phase at a flow rate of 1 mL/min. The results of the analysis are shown in In
addition, the degree of introduction of an oleoyl group in deuterated chloroform was analyzed
with a Bruker AVANCE DPX 400 1H nuclear magnetic resonance spectrometer. The
molecular weight of the cationic lipid synthesized under the ions of MeOH: 5 mM
ammonium formate-0/25% formic acid (70:30) was analyzed using t Technologies 646
Triple quad mass spectrometer. Through the above is, it was confirmed that the oleoyl
group was introduced to one end of the triethylenetetramine. The yield was 73.8%, and 1.1
equivalents of oleoyl groups were introduced into triethylenetetramine. Based on GPC, the
purity was confirmed to be 96.7%.
Example 2: Synthesis of 1,6-dioleoyl triethylenetetramide
0.50 g (3.34 mmol) of triethylenetetramine and 2.00 g (6.69 mmol) of methyl oleate
were placed in a round bottom flask and then allowed to react with stirring with a magnetic
bar at 65°C under nitrogen for 5 days.
After completion of the on, the process of adding 15 mL of hexane to the
reaction product to precipitate 1,6-dioleoyltriethylenetetramide and extracting unreacted
methyl oleate was repeated three times. The precipitated lipid was precipitated and separated
from hexane by centrifugation, recovered and vacuum dried.
The lar weight of the purified cationic lipid and the degree of introduction of
oleoyl groups were confirmed by gel chromatography, proton nuclear magnetic resonance
spectroscopy and mass spectrometry in the same manner as in Example 1. The results of the
gel chromatography and proton nuclear magnetic nce spectroscopy are shown in FIGS.
1 and 2, respectively. The yield was 79.9%, and 2.06 equivalents of oleoyl groups were
introduced into the triethylenetetramine. Based on GPC, the purity was med to be
95.7%.
Example 3: Synthesis of Trioleoyl triethylenetetramide
1,3,6-trioleoyl triethylenetetramide was synthesized by further reacting 1,6-dioleoyl
triethylenetetramide synthesized in Example 2 with methyl oleate. Specifically, 400 mg
(578.3 μmol) of 1,6-dioleoyl triethylenetetramide and 173.2 mg (578.3 μmol) of methyl oleate
were dissolved in 100 mL of dimethylformamide and then allowed to react with refluxing and
stirring at 90°C under nitrogen for 5 days.
After completion of the reaction, the reaction product was vacuum dried to remove
dimethylformamide, and then 50 mL of hexane was added to precipitate unreacted 1,6-
dioleoyl triethylenetetramide and then centrifuged. Subsequently, the separated supernatant
was vacuum dried, to which 10 mL of 1M hydrogen de (HCl) was added, and the
synthesized trioleoyl triethylenetetraamide was converted in the form of a mono-HCl
salt (1,3,6-trioleoyl triethylenetetramide·1HCl). After that, 50 mL of form was added
thereto and unreacted methyl oleoyl was extracted and d in a separating funnel. The
acidic aqueous on in which the cationic lipid was dissolved was neutralized with sodium
hydroxide, and the lipid was extracted with chloroform and vacuum dried.
The molecular weight of the purified and finally obtained product and the degree of
introduction of oleoyl groups were confirmed by using gel chromatography, proton nuclear
magnetic resonance spectroscopy and mass ometry in the same manner as in Example 1.
The results of the gel chromatography are shown in It was confirmed that the yield
was 47.5% and 2.94 equivalents of oleoyl groups were bonded to the triethylenetetramine.
Based on GPC, the purity was confirmed to be 94.3%.
Example 4: Synthesis of Tetraoleoyl triethylenetetramide
0.50 g (3.34 mmol) of triethylenetetramine and 8.00 g (26.76 mmol) of methyloleate
were placed in a round bottom flask and then allowed to react with stirring with a magnetic
bar at 65°C under nitrogen for 5 days.
After completion of the reaction, the process of adding 15 mL of hexane to the
reaction product to precipitate tetraoleoyl triethylenetetramide and extracting ted
methyloleate was repeated three times. The itated tetraoleoyl triethylenetetramide lipid
was precipitated and ted from hexane by centrifugation, recovered and vacuum dried.
The molecular weight of the purified cationic lipid and the degree of introduction of
oleoyl groups were confirmed by gel chromatography, proton nuclear magnetic resonance
spectroscopy and mass spectrometry in the same manner as in Example 1. The results of the
gel chromatography and proton nuclear magnetic resonance spectroscopy are shown in FIGS.
1 and 3, respectively. The yield was 89.1% and 4.05 lents of oleoyl groups were
introduced into the triethylenetetramine. Based on GPC, the purity was confirmed to be
99.4%.
【
Claims (1)
1.】 【Claim 1】 5 A method for preparing a cationic lipid represented by Formula 1, comprising reacting an oligoalkyleneamine represented by Formula 2, with a fatty acid alkyl ester represented by Formula 3 to prepare the cationic lipid of Formula 1: [Formula 1] 10 [Formula 2] [Formula 3] in the formulae 1 to 3, 15 n and m are independently 0 to 12, with the proviso that 1 ≤ n + m ≤ 12, a and b are independently 1 to 6, R1, R2, R3 and R4 are each independently hydrogen, or saturated or unsaturated fatty acid group having 12 to 26 carbon atoms, with the proviso that at least one of R1 and R4 is saturated or rated fatty acid group having 12 to 26 carbon atoms, 20 R is ted or unsaturated hydrocarbon having 11 to 25 carbon atoms, and R5 is an alkyl group having from 1 to 14 carbon atoms. 【Claim 2】 The method for preparing a cationic lipid according to claim 1, wherein the molar ratio of the oligoalkyleneamine to the fatty acid alkyl ester is adjusted to more than 1 to 20 or less to obtain the cationic lipid of Formula 1 where one of R1 and R4 is hydrogen and R2 and R3 are hydrogen. 【Claim 3】 The method for preparing a cationic lipid according to claim 1, wherein the molar 5 ratio of the fatty acid alkyl ester to the oligoalkyleneamine is adjusted to 1 or more to 5 or less to obtain the ic lipid of Formula 1 where R1 and R4 are independently saturated or unsaturated fatty acid groups having 12 to 26 carbon atoms and R2 and R3 are hydrogen. 【Claim 4】 The method for preparing a cationic lipid according to claim 1, wherein the molar 10 ratio of the fatty acid alkyl ester to the oligoalkyleneamine is adjusted to more than 5 to 20 or less to obtain the cationic lipid of Formula 1 where R1, R2, R3 and R4 are independently saturated or unsaturated fatty acid group having 12 to 26 carbon atoms. 【Claim 5】 A method for preparing a cationic lipid of Formula 1 of claim 1 where R1 and R4 are 15 ted or rated fatty acid group having 12 to 26 carbon atoms, and one of R2 and R3 is hydrogen, wherein the method comprises reacting the cationic lipid of Formula 1 defined in claim 1 where R1 and R4 are saturated or unsaturated fatty acid group having 12 to 26 carbon atoms and R2 and R3 are hydrogen, with a fatty acid alkyl ester of Formula 3 d in claim 1 to prepare the cationic 20 lipid of Formula 1 where R1 and R4 are saturated or unsaturated fatty acid group having 12 to 26 carbon atoms and one of R2 and R3 is hydrogen. 【Claim 6】 The method for preparing a cationic lipid according to claim 5, wherein the molar ratio of the fatty acid alkyl ester to the cationic lipid of Formula 1 is 0.5 or more to 20 or less. 25 【Claim 7】 The method for preparing a cationic lipid according to any one of claims 1 to 6, wherein n and m are ndently 0 to 9, with the proviso that 1 ≤ n + m ≤ 10. 【Claim 8】 The method for preparing a cationic lipid according to any one of claims 1 to 6, wherein n and m are independently 2 to 4. 【Claim 9】 The method for preparing a cationic lipid according to any one of claims 1 to 6, wherein one or more of R1, R2, R3, and R4 are independently unsaturated fatty acid group 5 having 14 to 22 carbon atoms. 【Claim 10】 The method for ing a cationic lipid according to any one of claims 1 to 6, wherein one or more of R1, R2, R3, and R4 are independently selected from the group consisting of lauroyl, myristoyl, palmitoyl, stearoyl, arachidoyl, behenoyl, eroyl, 10 cerotoyl, myristoleoyl, palmitoleoyl, sapienoyl, oleoyl, linoleoyl, arachidonoyl, eicosapentaenoyl, erucoyl, docosahexaenoyl, and cerotoyl. 【Claim 11】 The method for preparing a cationic lipid according to any one of claims 1 to 6, wherein the oligoalkyleneamine of Formula 2 is oligoethyleneamine. 15 【Claim 12】 The method for preparing a cationic lipid according to claim 11, wherein the oligoalkyleneamine is selected from the group consisting of diethylenetriamine, triethylenetetramine, tetraethylenepentamine, pentaethylenehexamine, hexaethyleneheptamine, heptaethyleneoctamine, hylenenonamine, nonaethylenedecamine, 20 decaethyleneundecamine, undecaethylenedodecamine, ethylenetridecamine and tridecaethylenetetradecamine. 【Claim 13】 The method for preparing a cationic lipid according to any one of claims 1 to 4, wherein the oligoalkyleneamine of Formula 2 and the fatty acid alkyl ester of a 3 are 25 reacted in the absence of an organic t. 【Claim 14】 The method for preparing a cationic lipid according to any one of claims 1 to 6, further comprising adding a nonpolar organic solvent after the reaction, itating and separating the cationic lipid from the unreacted materials to purify the cationic lipid. 【Claim 15】 The method for preparing a cationic lipid according to claim 14, wherein the nonpolar organic solvent is an alkane or ether having 4 to 12 carbon atoms. 【Claim 16】 5 The method for preparing a cationic lipid according to claim 5, r comprising dissolving the cationic lipid in a nonpolar organic t, adding an acid to separate the cationic lipid as an acid addition salt in the s layer from the organic solvent, neutralizing the separated cationic lipid, and extracting and purifying the neutralized cationic lipid with a nonpolar organic solvent. 【
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| NZ758732A NZ758732A (en) | 2015-12-30 | 2016-12-29 | Method for selectively synthesizing cationic lipids |
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| Application Number | Priority Date | Filing Date | Title |
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| KR10-2015-0189843 | 2015-12-30 |
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