DIBI, a 3-hydroxypyridin-4-one chelator iron-binding polymer with enhanced antimicrobial activity

Article abstract of DOI:10.1039/c8md00192h

Depriving microorganisms of bioavailable iron is a promising strategy for new anti-infective agents. The new, highly water-soluble, low molecular weight co-polymer DIBI was developed to selectively bind iron(iii) ions as a tris chelate and acts as a standalone anti-infective. Minimum inhibitory concentration (MIC) studies show DIBI is effective against representative reference strains for Gram-positive and Gram-negative bacteria S. aureus and A. baumannii, and the fungus C. albicans. Compared to the small molecule iron chelators, deferiprone and deferoxamine, DIBI outclassed these by factors of 100 to 1000 for inhibition of initial growth. DIBI and a series of related co-polymers (M_w of 2-9 kDa) were synthesized via reversible addition-fragmentation chain transfer (RAFT) polymerization of a chelating 3-hydroxypyridin-4-one (HPO) methacrylamide monomer and N-vinylpyrrolidone (NVP). Full incorporation of the HPO monomer into the co-polymers from the reaction solution was determined by ¹H NMR spectroscopy and ranged from 4.6 to 25.6 mol%. UV-vis spectroscopy showed that all the HPO in DIBI binds readily to iron(iii) in a tris chelate mode to the maximum theoretical iron(iii) binding capacity of the co-polymer. Chemical characterization including single crystal X-ray diffraction analyses of the O-benzyl protected and the functional HPO monomer are discussed. By design, DIBI is highly water soluble; the highest mass fraction in water tested was 70% w/w, without the need of organic co-solvents.

Full text of DOI:10.1039/c8md00192h

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RESEARCH ARTICLE
DIBI, a 3-hydroxypyridin-4-one chelator iron-
binding polymer with enhanced antimicrobial
activity†
Cite this: DOI: 10.1039/c8md00192h
ad
ad
a
b
M. Trisha C. Ang,‡ Roger Gumbau-Brisa,‡ David S. Allan, Robert McDonald,
b
ac
d
Michael J. Ferguson,
Bruce E. Holbein and Matthias Bierenstiel
*
Depriving microorganisms of bioavailable iron is a promising strategy for new anti-infective agents. The
new, highly water-soluble, low molecular weight co-polymer DIBI was developed to selectively bind ironIJIII)
ions as a tris chelate and acts as a standalone anti-infective. Minimum inhibitory concentration (MIC) stud-
ies show DIBI is effective against representative reference strains for Gram-positive and Gram-negative
bacteria S. aureus and A. baumannii, and the fungus C. albicans. Compared to the small molecule iron che-
lators, deferiprone and deferoxamine, DIBI outclassed these by factors of 100 to 1000 for inhibition of ini-
w
tial growth. DIBI and a series of related co-polymers (M of 2–9 kDa) were synthesized via reversible addi-
tion–fragmentation chain transfer (RAFT) polymerization of a chelating 3-hydroxypyridin-4-one (HPO)
methacrylamide monomer and N-vinylpyrrolidone (NVP). Full incorporation of the HPO monomer into the
1
co-polymers from the reaction solution was determined by H NMR spectroscopy and ranged from 4.6 to
Received 10th April 2018,
Accepted 12th June 2018
25.6 mol%. UV-vis spectroscopy showed that all the HPO in DIBI binds readily to ironIJIII) in a tris chelate
mode to the maximum theoretical ironIJIII) binding capacity of the co-polymer. Chemical characterization
including single crystal X-ray diffraction analyses of the O-benzyl protected and the functional HPO mono-
mer are discussed. By design, DIBI is highly water soluble; the highest mass fraction in water tested was
70% w/w, without the need of organic co-solvents.
DOI: 10.1039/c8md00192h
rsc.li/medchemcomm
6
,7
opment
to fight this threat has prompted researchers
Introduction
8,9
worldwide to explore new strategies to combat infections.
The widespread development of antibiotic resistance over re-
cent decades has become a global public health concern.
One approach to combatting antibiotic resistance is
through limiting iron availability to microbes to restrict their
1–4
1
0
Less than six months after the first identification in China of
the mcr-1 gene resistant to colistin, an antibiotic drug class
considered of last resort, a patient in the United States was re-
growth and help facilitate infection clearance. In prokary-
otic and eukaryotic genera iron has several irreplaceable
physiological roles within enzymes needed for DNA synthesis
and repair, and thus restricting bioavailable iron limits cell
5
portedly infected. We have reached the age in which the cur-
1
1
rent suite of antibiotics is no longer a viable and effective so-
lution in combatting antibiotic resistant infection. In
addition, the lack of new types of anti-infectives under devel-
growth. The innate immune response in humans against
invading microbial pathogens includes sequestration of iron
intracellularly to limit its availability to the microorgan-
1
2
isms. To counter this and other defense mechanisms, path-
ogenic microbes produce receptor/uptake systems to directly
access the body's iron carriers such as transferrin or produce
a
Chelation Partners Inc., 1411 Oxford St. Suite 369, Halifax, Nova Scotia, B3H
3
Z1, Canada
small, high affinity iron-chelators, siderophores, to harvest
b
X-Ray Crystallography Laboratory, Department of Chemistry, University of
13,14
iron from their environment.
These microbial chelators
Alberta, Edmonton, Alberta, T6G 2G2, Canada
c
have a wide range of chemical structures and present one or
more types of binding sites, e.g. hydroxamate, catecholate
and citrate. The structure and distribution of binding sites in
microbial chelators afford high specificity towards ironIJIII)
ions, commonly presenting binding constants in the range of
Department of Microbiology and Immunology, Dalhousie University, 5859
College St., Halifax, Nova Scotia, B3H 1X5, Canada
d
Department of Chemistry, Cape Breton University, 1250 Grand Lake Rd, Sydney,
Nova Scotia, B1P 6L2, Canada. E-mail: matthias_bierenstiel@cbu.ca;
Tel: +1 (902) 563 1391
†
Electronic supplementary information (ESI) available. CCDC 1812776 and
830658. For ESI and crystallographic data in CIF or other electronic format see
2
0–30
Kcomplex of 10
, but capable of reaching much higher
1
values such as that for enterobactin. Enterobactin,
tricatecholate chelator, is produced by E. coli and can bind
a
DOI: 10.1039/c8md00192h
These individuals contributed equally to this work.
‡
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an ironIJIII) ion with a binding constant of Kcomplex of 10 , the
MedChemComm
5
0
iron complexes by microbes is reduced, if not stopped
completely, thus limiting their growth while anti-infective
drugs and host clearance defense mechanisms take effect.
Another aspect is that folding of a polymeric chelator around
an ironIJIII) ion would create a larger spatial barrier between
the metal centres and bacterial siderophores on the exterior
of the folded polymer chain. Although iron-binding polymers
have great potential, the approaches published so far are
fraught with difficulties such as complicated, multi-step syn-
thesis, low yields, and often require co-solvents to improve
1
5
highest value recorded for a microbial siderophore.
A potential strategy to deprive microbes of iron is to use
other ironIJIII) chelators to out-compete microbial
siderophores and acquisition systems in vivo. Deferoxamine
3
1
(
DFO, Kcomplex 10 ) and deferiprone (DFP) are two examples
of ironIJIII) chelators approved by the FDA and EMA for the
1
6–18
treatment of iron overload disorders in humans (Fig. 1).
DFO is a hexadentate hydroxamate-based chelator of bacterial
origin that has been shown to facilitate iron uptake by a wide
spectrum of bacteria, hence limiting its suitability as an anti-
2
5,34–36
solubility in water.
1
9
infective agent candidate to deprive bacteria of iron. DFP is
a fully synthetic HPO chelator that has been used to treat
thalassemia major for over two decades. This bidentate chela-
tor forms octahedral tris complexes with ironIJIII) and has a
The work in our group focuses on developing iron-binding
polymeric platforms with antimicrobial activity that follows a
streamlined, cost effective and scalable approach. In previous
publications we showed the efficacy of an iron-binding poly-
3
6 20–22
29,37–39
binding constant of Kcomplex of 10 .
The large complexa-
mer DIBI against microorganisms and cancer cells
and
4
0,41
tion constant puts DFP at par with naturally occurring micro-
bial siderophores, and its clinical applications for the treat-
ment of iron overload conditions make this chelator a very
interesting candidate to out-compete microbial siderophores
for iron. In vitro studies have shown that some bacteria are
inhibited by DFP but at very high chelator concentra-
its lack of toxicity in murine models.
The synthesis, basic
characterization, and broad-based antimicrobial activity of
DIBI, highly water-soluble, HPO containing poly-
vinylpyrrolidone co-polymer is described herein with addi-
a
3
0
tional detail as provided in a previous patent disclosure.
2
3,24
tions.
Other small iron chelators that incorporate the
Results and discussion
Iron-binding monomer in DIBI
HPO binding moieties have been synthesized with the aim of
competing against bacterial siderophores; however, among
the several iron chelators reported, only a few candidates
yielded moderate to low inhibitory activity against test
To maximize biocompatibility and confer iron-binding capa-
bility to DIBI we chose the HPO chelator motif. We
derivatized the hydroxypyridinone ring by introducing an
N-ethyl-2-methylacrylamide tether to enable olefin polymeriza-
tion, resulting in the monomer N-[2-(3-hydroxy-2-methyl-4-
oxopyridin-1IJ4H)-yl)ethyl]methacrylamide (5) (Scheme 1). The
synthesis of 5 was a modified procedure from that previously
reported by Feng et al. that uses a methacrylamide and
circumvented the installation of ethylenediamine before the
addition of an acrolein group (see ESI†). The first step in the
synthetic route of 5 is the formation of 3-benzyloxy-2-methyl-
4-pyrone (2) using benzyl chloride (BnCl) to protect the hy-
droxyl group. The next step is the addition of N-(2-
aminoethyl)-2-methyl-2-propeneamide (3) under basic aque-
ous conditions to generate the pyridin-4-one ring system of 4,
followed by the removal of the benzyl protecting group in
concentrated hydrochloric acid affording 5 as a beige-
coloured, crystalline powder.
2
5–29
microorganisms.
If iron sequestration from microbes is to become a feasi-
ble clinical strategy for the treatment of infections the chela-
tors used for this purpose must have a high affinity for
ironIJIII) ions, sequester iron in a way that is not suitable for
microbial uptake, and provide higher anti-microbial activity
than the currently employed clinical iron chelators. Addition-
ally, such chelators need to be water-soluble, possess low tox-
icity to humans, and have a simple synthetic preparation to
be economically viable.
2
7
We have developed a different approach to capture ironIJIII)
by using polymers containing iron binding sites rather than
3
0
small ligands. Ideally, such an iron-binding polymer should
have a high selectivity and capacity for iron and be structur-
ally unrecognizable as a suitable source of iron by microbes.
Due to their size, high molecular weight polymers would of-
fer low toxicity relative to low molecular weight molecules as
The molecular structure of the intermediate N-[2-(3-
(benzyloxy)-2-methyl-4-oxopyridin-1IJ4H)-yl)ethyl]-
methacrylamide (4) was obtained by X-ray crystallography
3
1–33
they would not be readily taken up by cells.
Transport of
(
Fig. S1†). Single crystals of 4 were obtained by slow evapora-
tion of an aqueous solution at room temperature. 4 crystal-
lized in the space group P212121 presenting four molecules re-
lated by symmetry in the unit cell. The X-ray crystal structure
shows the benzyl group and the N-ethyl-2-methacrylamide
tether to the nitrogen site of the HPO ring. The only hydro-
gen bonding interactions present in the crystal structure are
between the amide N–H bonds and adjacent HPO carbonyls
that propagate along the crystallographic axis b of the unit
cell (Fig. S2†).
Fig.
disorders.
1 FDA and EMA-approved iron chelators for iron-overload
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Scheme 1 Synthesis of 5 from maltol (1). (a) 1.1 equiv. BnCl, 1.05 equiv. NaOH, MeOH, 85 °C, 2 h; (b) MeOH : H
2
O (1 : 1), 2 N NaOH, 95 °C, 2 h; (c)
conc. HCl, r.t. overnight.
The molecular structure of the functional monomer N-[2-
3-hydroxy-2-methyl-4-oxopyridin-1IJ4H)-yl)ethyl]-2-methylprop-
Novel family of polyvinylpyrrolidone co-polymers
(
A family of co-polymers was conceptually designed as simple,
siderophore-inspired biocompatible polymers with high water
solubility. Polyvinylpyrrolidone (PVP) was chosen as the main
structural component for our iron binding co-polymers, as it
is highly soluble in water, and well known in the medical
field having been used as a plasma volume expander and as
an excipient ingredient in many pharmaceutical prepara-
2
-enamide (5) was also obtained by X-ray crystallography
(
Fig. 2). Single crystals were obtained by slow evaporation in
RPMI culture medium at 4 °C. The molecular structure shows
the free monomer with the carbonyl and hydroxyl group ac-
cessible for iron coordination. The water molecule is hydro-
gen bonding between three molecules of 5 via the carbonyl
groups and NH bond of the acrolein portion of 5. The hydro-
gen bonding network of 5 is propagated by pairs of carbonyl
and hydroxyl substituents of the ring (Fig. S3†).
42
tions. The polymerization of co-polymers was carried out
with varying ratios of 5 and commercially available NVP via
RAFT in mild aqueous conditions (Scheme 2 and Table 1).
The total monomer-to-RAFT agent ratio was held constant at
a ratio of 50 : 1.
The results in Table 1 show that the polymer content of
monomer 5 can be controlled by changing the ratio 5 : NVP
in the reaction solution. The highest mol% content of 5 gave
the lowest isolated yield of 55% that indicates difficulty to in-
corporate amounts higher than 15 mol% of 5. The RAFT me-
diated polymerization was found to provide a relatively nar-
row molecular weight distribution of the co-polymer products
The UV-vis characterization of 5, dissolved in water,
showed a band at 282 nm and two bands in the range 200–
2
40 nm (Fig. S4†). The signal at longer wavelengths corre-
sponds to the π–π* transition of HPO ring, whereas the ab-
sorption bands at shorter wavelengths correspond to the
overlapped contribution of the HPO ring and the
methacrolein moiety.
w
with the average M of the compositions being somewhat
similar at the different 5 : NVP conditions (see ESI†). Based
on these results it appeared that a 5 : NVP ratio of between 1 :
1
0 and 1 : 15 provided the best overall 5 content (10–15
mol%) while retaining high product yields and a PDI below
.0.
2
The evolved target polymer DIBI: synthesis and
characterization
We decided to further develop a co-polymer containing 15
mol% of 5 to maximize both iron binding capacity and poly-
merization yield. Our lead compound DIBI was obtained fol-
lowing the above synthesis as an off-white powder in 85%
yield. After 2 hours of reaction time the full incorporation of
5
from the reaction solution into DIBI was determined by fol-
1
lowing the suppression of the CH acrolein signals using H
NMR spectroscopy (see ESI†). The structure of DIBI com-
prises 15 mol% of 5, with PVP accounting for the rest of the
co-polymer. GPC characterization for DIBI determined a Mw
of 9 kDa with a polydispersity index of 1.2 (see ESI†). The
good correlation between refractive index and UV-vis profiles
indicated a homogeneous composition of 5 across the
Fig. 2 Molecular structure of 5. Displacement ellipsoids for non-H
atoms are shown at 30% probability level and H-atoms are represented
by circles of arbitrary size.
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Scheme 2 Synthesis of DIBI (M
n
: 7.5 kDa, M
w
: 9 kDa, PDI: 1.2); TBHP = tert-butyl hydroperoxide, TMEDA = N,N,N′,N′-tetramethylethylenediamine,
and RAFT-agent = 2-[(ethoxymethanethioyl)sulfanyl]-2-methylpropanoic acid.
a
Table 1 Characterization data of a series of co-polymers with varying ratio 5: NVP
b
c
d
Co-polymer
5 : NVP in reaction solution
Content of 5 in polymer (mol%)
Yield
M
w
PDI
a
b
c
1 : 20
1 : 15
1 : 10
1 : 5
4.6
9.5
15.3
25.6
88%
80%
85%
55%
3656
4799
3430
2298
1.7
1.9
1.6
1.4
d
a
b
1
c
Total monomer content of 50 equivalents relative to remainder of reagents. Measured using H NMR spectroscopy (see ESI). Relative
d
w n
molecular weight. Polydispersity index calculated as M /M .
distribution of polymer molecular weights. By design, DIBI is
extremely water-soluble; aqueous solutions of DIBI of 20% w/
w were routinely used in our work. The highest mass fraction
in water tested was 70% w/w, without using organic solvents
also determined in MOPS buffer at pH 7.42 by interpolating
the intensity of the HPO signal of DIBI in a standard curve of
5 in the same conditions. The results indicated an average
content of 13 mol% or 24 mass% of 5 in DIBI, in good agree-
1
(e.g. DMSO) to improve polymer solubility. In comparison,
ment with the mol% content of 5 determined by H NMR
DFP and DFO (mesylate salt) have a maximum water solubil-
ity of 1.4% w/w and 5% w/w, respectively.
spectroscopy (see ESI†).
The UV-vis spectrum of DIBI in water shows the expected
HPO band with maximum at 282 nm, as observed in 5, and a
group of overlapped signals below 230 nm formed by PVP
and amide signals of 5 (Fig. S5†). The extinction coefficient
of DIBI is about 9.5 times that of 5 that correlates an average
of 9.5 moles 5 per mol of DIBI. The mol% of 5 in DIBI was
Iron-binding capacity of DIBI
The iron-binding activity of DIBI is provided by its platform
supported HPO in 5 which are widely distributed within the
linear structure of DIBI. The bidentate nature of the chelating
monomer 5 led us to expect the formation of octahedral tris
Fig. 3 UV-vis spectra showing the MLCT band for the ironIJIII) titration of 122 mM DIBI at pH 7.42 in MOPS. Inset: Plot of absorbance maxima at
60 nm vs. iron concentration for the ironIJIII) titration.
4
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Table 2 Minimum inhibitory concentrations for S. aureus (ATCC 43300)
at 48 h for co-polymers, PVP, 5 and DFP in RPMI
of a structural enhancement of antibacterial activity by the
incorporation of 5 into a polymer. The results also indicated
lower MICs were achieved by increasing the amount of 5 in-
corporated into the co-polymer. The control experiment for
MIC
Content of 5 in
polymer (mol%)
a
−1
b
c
Co-polymer
μg mL
μM
μMFe
PVP homopolymer (10 kDa M ) showed no growth inhibition
w
a
b
c
PVP
5
9.5
8
4
2
0.21
0.13
0.15
25 600
530
0.07
0.04
0.05
N/A
176
96
even at very high concentrations. RPMI was chosen as the as-
say medium as it is more representative of the host environ-
ment compared to other media choices with regard to its
ironIJIII) content (approx. 0.5 μM).
15.3
25.6
0
N/A
N/A
256 000
125
40
DFP
288
a
1
b
Measured using
H
NMR spectroscopy (see ESI).
Molarity
expressed as equivalent concentration of 5 based on the mol% of 5
in each polymer. MIC value normalized for maximum amount of
Antimicrobial activity of DIBI and other medical ironIJIII) chelators
c
Table 3 shows the MIC for DIBI, DFP and DFO against ATCC
reference strains of S. aureus and A. baumannii and C.
albicans. These opportunistic pathogens were chosen as rep-
resentative Gram-positive/negative bacteria and yeast, respec-
tively. S. aureus and A. baumannii have been highlighted re-
cently by the WHO as priority targets for the development of
new antibacterial treatments because they pose a serious
iron bound by each chelator, assuming octahedral coordination with
the following (ligand : Fe) ratios: co-polymer (3 : 1), 5 (3 : 1), DFP (3 :
1). The ratio for co-polymers is expressed according to the tris-
chelate complexes of 5.
complexes with ironIJIII), as seen in other HPO coordination
3
6,43–45
1
complexes.
Thus, based on NMR and UV-vis characteri-
threat to human health in community and clinical settings.
zation the theoretical chelating capacity of a 122 mM solution
of DIBI would be equivalent to 1120 μM of 5 and 373 μM of
ironIJIII) for the expected DIBI–Fe complex. The electronic
spectra obtained from the batch titration of DIBI solutions
ironIJIII) nitrate showed a metal-to-ligand charge transfer
C. albicans is one of the most common fungal yeast opportu-
nistic pathogens in hospital-acquired infections due to their
4
6
rapid growth on medical devices or on human tissue.
Table 3 shows the measured MIC values for DIBI, DFP
and DFO in different units to compare them to anti-infectives
−
1
(MLCT) band at 460 nm, indicative of the formation of the
(μg mL ) and other chelators (μM) in the literature. The
values expressed in μMFe are normalized based on the
ironIJIII) complex stoichiometry for each chelator, assuming
the formation of the octahedral complexes for each ligand.
For DIBI the μMFe values are expressed as the concentration
of 5 based on 24% mass content.
ironIJIII) complex with DIBI (Fig. 3). Plotting the absorption
maxima at 460 nm against ironIJIII) concentration gave a pla-
teau at 371 μM ironIJIII). This indicated that all binding sites
of 5 within DIBI are available for the tris-chelation of iron,
resulting in a total iron-binding capacity of 338 μmol of iron
per gram of DIBI. No precipitate was visible during the iron
titration experiments, and the coloured films obtained upon
solution evaporation re-dissolved readily following weeks of
being dry.
The MIC results obtained indicated that DIBI inhibited all
4
7
three classes of microbes displaying very low MICs. The MIC
values of DIBI were 2 and 3 orders of magnitude lower than
the small molecule iron chelators, which require concentra-
−1
tions in the mg mL range to achieve even modest antimicro-
bial activity. DFP was more effective at inhibiting bacterial
growth over that of fungal yeast C. albicans, whereas DFO was
non-inhibitory for all organisms at the maximum concentra-
tion tested. Control experiments using commercially available
Antimicrobial activity of the family of co-polymers
Select co-polymers presented in Table 1 with relative molecu-
lar weights in the range 2–4 kDa were tested using MIC to as-
sess their activity against the growth of S. aureus in RPMI,
shown in Table 2. The co-polymers demonstrated extremely
low MIC values for S. aureus compared to the free monomer
PVP (M 10 kDa) showed no antimicrobial effect.
w
Consistent with our results, it has been reported that the
presence of DFO, a bacterial siderophore itself, in culture me-
dia can improve microbial growth in vitro while DFP can be
5
and the clinical iron chelator DFP. This is strong evidence
Table 3 Minimum inhibitory concentrations at 48 h for DFO, DFP and DIBI in RPMI
MIC
DIBI
DFP
DFO
−1
b
c
−1
c
−1
c
Microorganism
μg mL
μM
μMFe
μg mL
μM
288
575
1150
μMFe
μg mL
μM
μMFe
a
S. aureus ATCC 43300
A. baumannii ATCC 19606
C. albicans ATCC SC5314
4
2
2
4.06
2.03
2.03
1.35
0.67
0.67
40
80
160
96
192
383
>1280
>1280
>1280
>1949
>1949
>1949
>1949
>1949
>1949
a
b
c
Methicillin-resistant Staphylococcus aureus (MRSA). Concentration of 5 based on 24 mol% content in DIBI. MIC value normalized for
maximum amount of iron bound by each chelator, assuming octahedral coordination with the following (ligand : Fe) ratios: DFO (1 : 1), DFP
(3 : 1), DIBI (3 : 1). The ratio for DIBI is expressed according to the tris-chelate complexes of 5.
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1
9,23,48
mildly inhibitory.
However, the internalization of large
tre for access to equipment. The assistance of Maria Parquet
with MIC determinations is also acknowledged.
molecular size polymers or polymer-bound iron by bacteria is
less likely because specialized, active transport mechanisms
are needed over 600 Da.
1
4
References
We hypothesize that the enhanced antimicrobial efficacy
shown by DIBI can be attributed to its polymer structure.
Given the structural similarities between binding sites in DFP
and 5, it is reasonable to consider that the complexation con-
stant for 5 would be, at best, of the same magnitude as for
DFP. This leads to the possibility that having 5 tethered and
dispersed throughout one polymer backbone results in en-
hanced iron-binding due to intramolecular proximity of
neighbouring 5 binding moieties. Therefore, the resulting lo-
cal concentration of 5 within the polymer platform would be
effectively higher than for the free monomers at an equiva-
lent concentration in solution. An additional benefit of hav-
ing a polymeric structure is the potential for spatial hin-
drance, which may allow DIBI to sequester ironIJIII) ions away
from smaller siderophores.
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Conclusions
The novel water-soluble co-polymer DIBI displays low MICs
in vitro against the bacteria S. aureus and A. baumannii, and
the fungus C. albicans. Compared to the low molecular
weight clinically used iron chelators, DFO and DFP, DIBI
displayed 100 to 1000 times lower MICs against three classes
of microbes. DIBI is one of a series of co-polymers reported
that were synthesized from the monomers 5 and NVP using
RAFT polymerization. The antibacterial activity of DIBI and
related co-polymers stems from the amount of HPO-based
ironIJIII)-binding moieties of 5 present in their composition,
which can be easily controlled based on the 5 : NVP feed ratio
for the co-polymerization. It is hypothesized that the en-
hanced antimicrobial activity of DIBI compared to 5 is due to
the inability of microbes to internalize the polymer and to
the intramolecular proximity of neighbouring 5 binding moi-
eties. The possibility of synergistic effects between DIBI and
antibiotics against microbial pathogens is currently under
5 P. McGann, E. Snesrud, R. Maybank, B. Corey, A. C. Ong, R.
Clifford, M. Hinkle, T. Whitman, E. Lesho and K. E.
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