Measuring Rate and Equilibrium Constants of ®²¢â‚¬¯Cyclodextrin...

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Transcript of Measuring Rate and Equilibrium Constants of ®²¢â‚¬¯Cyclodextrin...

  • Measuring
Rate
and
Equilibrium
Constants
of
β‐Cyclodextrin‐Small
Molecule
Drug



    Non‐Covalent
Interactions
with
Capillary
Electrophoresis



    By


    Jennifer
Logie


    Honors
thesis
submitted
to
the
Faculty
of
Science
Department
of
Chemistry
 In
partial
fulfillment
of
the
requirements
of
the
B.
Sc.
Degree


    Supervisor:
Maxim
Berezovski


    Department
of
Chemistry


    University
of
Ottawa


    Ottawa,
Canada


    ©
April
2011,
Jennifer
Logie


  • 
 ii


    Abstract


    Cyclodextrin
is
a
circular
oligosaccharide
made
of
6‐8
glucose
monomers,
giving
the
ideal


    conformation
to
form
inclusion
complexes
with
many
different
small
molecules.
In
the


    pharmaceutical
industry,
it
is
commonly
used
as
an
excipient
to
increase
the
solubility
of
small


    molecule
drugs.
The
purpose
of
this
research
was
to
measure
equilibrium
and
rate
constants


    between
small
molecules
and
cyclodextrins
using
the
Equilibrium
Capillary
Electrophoresis
of


    Equilibrium
Mixtures
(ECEEM)
method.
In
ECEEM,
an
equilibrium
mixture
of
small
molecule,


    cyclodextrin
and
complex
is
injected
into
a
capillary
and
subjected
to
electrophoretic
separation.


    The
running
buffer
contains
the
same
concentration
of
cyclodextrin
as
in
the
equilibrium
mixture.


    Binding
parameters
of
complexes
can
be
found
by
using
the
time
propagation
pattern
and
shapes


    of
the
peaks
obtained
when
the
concentration
of
cyclodextrin
is
gradually
increased.
In
this
study,


    eight
small
molecule
drugs:
ibuprofen,
s‐flurbiprofen,
4,4’‐(propane‐1,3‐diyl)
dibenzoic
acid,


    resveratrol,
naproxen,
diclofenac,
folic
acid
and
phenylbutazone
were
studied
for
their


    complexation
with
β‐cyclodextrin.

The
ECEEM
method
proved
to
be
a
valuable
technique
for


    determining
the
practicality
of
cyclodextrin
as
a
drug
delivery
system
for
specific
drugs.




  • 
 iii


    Acknowledgements


    Without
the
help
and
support
of
the
Berezovski
lab,
this
work
could
not
have
been
completed.


    Particularly
the
supervision,
guidance
and
insight
of
Dr.
Maxim
Berezovski;
the
math
skills
of
Dr.


    Victor
Okhonin;
and
finally
the
instruction
and
help
of
Gleb
Mironov.



  • 
 iv


    Table
of
Contents


    List
of
Figures
......................................................................................................................................
v


    Statement
of
Contribution
................................................................................................................
vi


    1.
Introduction
....................................................................................................................................
1
 1.1
Rationale
.................................................................................................................................................
1
 1.2
Cyclodextrin
as
a
Drug
Delivery
Agent
....................................................................................................
2
 1.3
Binding
Constants
of
Cyclodextrin‐Small
Molecule
Complexes
..............................................................
5
 1.4
Literature
Review
of
KD
Determination
Methods
...................................................................................
6
 1.5
Principles
of
Capillary
Electrophoresis
....................................................................................................
9


    2.
Results
and
Discussion
..................................................................................................................
13
 2.1
4,4’
(Propane‐1,3
Diyl)
Dibenzoic
Acid
Model
......................................................................................
13
 2.2
Multiplex
Application
............................................................................................................................
23


    3.
References
....................................................................................................................................
31


    4.Experimental
..................................................................................................................................
33
 4.1
Chemicals
and
Materials
.......................................................................................................................
33
 4.2
Experimental
Conditions
.......................................................................................................................
34


    
 


  • 
 v


    List
of
Figures


    1.
Structures
of
Small
Molecule
Drugs
................................................................................................
3


    2.
β‐cyclodextrin
Structure
and
Inclusion
Complexation
....................................................................
4


    3.
Schematic
of
ECEEM
Set‐up
..........................................................................................................
11


    4.
Limit
of
Detection
of
PDDA
Electropherogram
.............................................................................
15


    5.
Degradation
of
PDDA
Electropherogram
......................................................................................
17


    6.
PDDA
with
varying
β‐cyclodextrin
concentrations
Electropherogram
.........................................
18


    7.
Velocity
of
Equilibrium
Mixture
as
a
Function
of
β‐cyclodextrin
Concentration
..........................
20


    8.
ECEEM‐MS
Schematic
...................................................................................................................
22


    9.
3‐D
Absorption
Plots
of
Small
Molecules
......................................................................................
24


    10.
2‐D
and
3‐D
Absorption
Plots
of
Multiplex
.................................................................................
25


    11.
Multiplex
with
varying
β‐cyclodextrin
concentrations
Electropherogram
.................................
27


    12.
Multiplex
KD
determination
Graphs

...........................................................................................
29
 


  • 
 vi


    Statement
of
Contribution


    Dr.
Serge
Gorelsky
did
the
DFT
calculations
to
determine
the
model
of
the
inclusion
complex


    between
β‐cyclodextrin
and
ibuprofen.
Victor
Okhonin
created
the
mathematical
model
used
to


    calculate
binding
parameters
for
the
PDDA‐β‐cyclodextrin
inclusion
complex.
Gleb
Mironov
did
all


    CE‐MS
work
that
is
referenced
in
Section
2.



  • 
 1


    1.
Introduction


    1.1
Rationale


    Pharmaceutical
companies
spend
billions
of
dollars
each
year
researching
new
and
exciting
drug


    candidates.
Getting
these
drugs
into
the
body
is
an
ongoing
battle
due
to
the
often‐low
solubility


    of
these
molecules.
Often
drugs
are
formulated
with
excipients,
which
use
various
methods
to


    increase
the
bioavailability
of
the
drugs
(1).
In
this
study,
several
common
drugs
on
the
market
will


    be
used
to
create
a
model
for
the
evaluation
of
drug
delivery
systems.




    Some
of
the
oldest
and
most
commonly
used
drugs
available
are
the
non‐steroidal
anti‐

    inflammatory
class
(NSAIDS).
Although
the
structures
of
these
drugs
vary,
th