Development of synthetic methodologies for ion channels
Date
2018-11-08
Authors
Hu, Chi-wei
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Abstract
The goal of this research was to develop a synthetic strategy to synthesize voltage-gated
(end-differentiated) ion channels with a minimal synthetic effort and this goal was
pursued in two different ways with two different structural components: macrocycles and
non-macrocycles (flexible acyclic components).
This thesis started with an end-differentiated macrocycle differentially protected with Boc
and nitro as amine protecting groups. To test the macrocycle’s suitability as a part of an
ion channel, a centrosymmetric ion channel candidate was synthesized. The synthesis
started with the reduction of the nitro to a free amine followed by a dimerization reaction
with terephthaloyl dichloride to form a centrosymmetric diamide. The Boc protecting
groups were removed to yield the target. Although the target compound showed ion
channel activity, its very poor solubility made it almost impossible for a complete
property investigation.
The second part of this research was to examine the necessity of macrocycles for ion
channels. Some centrosymmetric non-macrocyclic bolaamphiphiles related to a known
macrocyclic ion channel were synthesized and tested for their ion transport properties. An
oligoester was prepared from 2-[2-(2-chloroethoxy)ethoxy]ethanol and dodecanedioyl
dichloride followed by estérification with the mono octyl ester of maleic acid to yield a
diene that reacted with mercaptoacetic acid to give the target compound. An oligoester
homolog was also synthesized via the same chemistry.
Other compounds with ester-amide and amide functionalities were also synthesized.
The ester-amide compounds were synthesized from a mono Boc protected 1,8 -diamino-
3,6-dioxaoctane followed by acylation with dodecanedioyl dichloride to yield a diamide
that further reacted with the mono octyl ester of fumaric acid to afford a diene. The diene
reacted with mercaptoacetic acid to give the target ester-amide compound. The acyclic
hexamide compound used the diamide intermediate above to react with the mono Nmethyl-
N-octyl amide of maleic acid to afford a diene that reacted with mercaptoacetic
acid to give the target compound. The other ester-amide compound started with 1,8-
bis(methylamino)-3,6-dioxaoctane and followed the chemistry for the previous esteramide
to yield the target compound.
All the acyclic compounds were tested for their ion transport properties by pH-stat and
carboxyfluorescein release experiments. Among the compounds tested, oligoesters
showed clear ion channel activity, implying macrocycles are not necessary for ion
channel formation. On the other hand, ester-amides were found to be active only in
concentration ranges where they form large membrane defects. No ion transport activity
was found for the hexamide compound.
The third part of this thesis employed solid-phase synthesis to prepare some acyclic
oligoesters as ion channel candidates. The synthesis provides end-differentiated
compounds as required for voltage-gating applications. Building blocks with an acid
functional group and THP or TBDMS protected alcohol were prepared for the solid-phase
synthesis. Sequential coupling on Wang resin followed by cleavage and gel
filtration gave products that showed the expected NMR spectra. The MALDI mass
spectrum and GPC showed these samples contain deletion sequences due to incomplete
conversion. The coupling efficiency was calculated be an average 93% for each step. The
products formed active ion channels in a planar bilayer experiment, implying that this
synthesis has achieved the goal of the thesis.
Description
Keywords
Ion channels, Research, Methodology