A water soluble macromolecular nanobox having porphyrinic walls as a large host for giant guests

Article abstract of DOI:10.1002/pola.26515

Cyclic tetra{5,15-di-[p(ω-methoxypolyethyleneoxy)phenyl]-10,20-[p- oxyphenyl] methylen porphyrin}, cy-[O-(H₂-PTPEG₂)-O- CH₂-]₄, a water soluble macromolecule consisting of four porphyrin units [each with two long ω-methoxypolyethyleneoxy (PEG) branches bound on its peripheral positions] linked by means of four methylenoxy bridges, was prepared by an interfacial etherification reaction. Structural and spectroscopic characterization of cy-[O-(H₂-PTPEG₂)-O- CH₂-]₄ and of its cobalt-derivative {cy-[O-(Co-PTPEG ₂)-O-CH₂-]₄} was performed by means of MALDI-TOF mass spectrometry, NMR, UV-vis, and circular dichroism spectroscopy. The data obtained from the cy-[O-(Co-PTPEG₂)-O-CH₂-] ₄/Gramicidin-S mixture showed that some evident spectral changes were compatible with the formation of a supramolecular structure between the porphyrinic nanobox and the Gramicidin S (a polypeptide having a relevant pharmacological importance). These preliminary data highlight how cy-[O-(H ₂-PTPEG₂)-O-CH₂-]₄ and/or its metalled derivatives, for their both chemical composition and structural arrangement, have promising properties for applications as a drug carrier in aqueous media.

Full text of DOI:10.1002/pola.26515

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A Water Soluble Macromolecular Nanobox Having Porphyrinic
Walls as a Large Host for Giant Guests
Placido Mineo,1,2 Fabiola Spitaleri,1 Emilio Scamporrino1
1
ꢀ
Dipartimento di Scienze Chimiche and I.N.S.T.M. UdR of Catania, Universita di Catania, Viale A. Doria, 6, 95125 Catania, Italy
2CNR-IPCF Istituto per i Processi Chimico Fisici, Viale Ferdinando Stagno D’Alcontres, 37, 98158 Messina, Italy
Correspondence to: P. Mineo (E-mail: gmineo@unict.it)
Received 1 October 2012; accepted 28 November 2012; published online 7 January 2013
DOI: 10.1002/pola.26515
ABSTRACT: Cyclic tetra{5,15-di-[p(x-methoxypolyethyleneoxy)-
phenyl]-10,20-[p-oxyphenyl] methylen porphyrin}, cy-[O-(H2-
PTPEG2)-O-CH2-]4, a water soluble macromolecule consisting
of four porphyrin units [each with two long x-methoxypolye-
thyleneoxy (PEG) branches bound on its peripheral positions]
linked by means of four methylenoxy bridges, was prepared
by an interfacial etherification reaction. Structural and spectro-
scopic characterization of cy-[O-(H2-PTPEG2)-O-CH2-]4 and of its
cobalt-derivative {cy-[O-(Co-PTPEG2)-O-CH2-]4} was performed
by means of MALDI-TOF mass spectrometry, NMR, UV–vis,
and circular dichroism spectroscopy. The data obtained from
the cy-[O-(Co-PTPEG2)-O-CH2-]4/Gramicidin-S mixture showed
that some evident spectral changes were compatible with the
formation of a supramolecular structure between the porphyr-
inic nanobox and the Gramicidin S (a polypeptide having a rel-
evant pharmacological importance). These preliminary data
highlight how cy-[O-(H2-PTPEG2)-O-CH2-]4 and/or its metalled
derivatives, for their both chemical composition and structural
arrangement, have promising properties for applications as a
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drug carrier in aqueous media.
2013 Wiley Periodicals, Inc.
J. Polym. Sci., Part A: Polym. Chem. 2013, 51, 1428–1435
KEYWORDS: host-guest systems; hydrophilic polymers; MALDI;
polyethers; sensors; star polymers
INTRODUCTION In the last years, the biomedical research
has had to deal with complex problems related to the
increased incidence of cancerous pathologies as a conse-
quence of an increment of the average life and of the exposi-
tion to environments increasingly polluted.
acute or chronic toxicity so that alternative more functional
strategies are searched.
All this has led to the development of new molecular sys-
tems that can: (i) recognize the target molecules; (ii) be, in
turn, a potential drug; (iii) be analytically determinable; and
(iv) carry specific drugs, releasing them only where
necessary.
Efforts in this field are pharmacological oriented towards the
development of ‘‘smart molecules’’ able to selectively recog-
nize the diseased cells on which to act.1a The strategies until
now followed concern the use of molecules with suitable
cavities in their structure where to insert the drug molecules
with pharmacological activity.1b
For their peculiar properties, excellent candidates to satisfy
the first three points are the porphyrin compounds. How-
ever, as a negative aspect, the marked hydrophobic nature of
the heteromacrocycle core is, unfortunately, responsible of a
strong insolubility of these compounds in aqueous media. In
some cases, by the inclusion of ionic groups in the porphyrin
structure the insolubility has been mitigated6–8 without sub-
stantially change the affinity for the DNA molecules, but of-
ten the electric interaction with the cell membranes prevents
their crossing.9
Generally, for larger drug molecules vesicular systems are
used. They are often complex aggregates systems consisting
of amphiphilic species that tend to form, in aqueous environ-
ment, micellar structures in which the active drug molecules
may be trapped.2–5
Unfortunately, with these systems, the drug is however,
brought indiscriminately throughout the body, and released,
often without any specific control, in all tissues and also in
healthy cells. Moreover, the amphiphilic material (present in
large excess with respect to the amount of the drug) is
metabolized by the body with the possible occurrence of an
Recently, some of these water-soluble porphyrins have been
tested in important applications such as the treatment of
transmissible spongiform encephalopathy (known as the bo-
vine variant ‘‘mad cow disease’’), in chemotherapy and in
photodynamic therapy.10–13
Additional Supporting Information may be found in the online version of this article.
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However, another problem concerns the drug release from
the porphyrin complexes. In fact, porphyrin systems and, in
particular, their metal derivatives, easily form supramolecu-
lar complexes with suitable electron-rich bio-molecules (as
amino acids and drugs) but more complicated is their con-
trolled release and only a few studies concerning this prob-
lem are in literature.
matrix. Mass spectrometer calibration and average molecular
mass determination were performed as reported in previous
cases.17–19
Each m/z value reported in the spectra and in the text is
referred to the ion containing the most abundant isotope of
each element present in the molecule.
Synthesis of Tetrakis-(p-hydroxyphenyl)porphyrin,
H2-P(OH)4
This product was prepared according to the method
described by Little et al.,20 starting from pyrrole and p-ace-
toxybenzaldehyde in boiling propionic acid.
Recently, as a first step of a work in this topic, we have syn-
thesized some cyclic formal-porphyrin oligomers,14 soluble in
organic solvents. Now, in this work we have designed a simple
and fast procedure to synthesize uncharged water soluble
macromolecular nanobox. In particular, we report the synthe-
sis of cyclic tetra{5,15-di-[p(x-methoxypolyethyleneoxy) phe-
nyl]-10,20-[p-oxyphenyl]methylen porphyrin}, (indicated in
the followed as cy-[O-(H2-PTPEG2)-O-CH2-]4) which, having a
3D structure with four porphyrin units held together by ether
formal bridges, forms as a molecular box. Besides the synthe-
sis, chemical and spectroscopic characterization of this cyclic
tetra-porphyrin-ether (and its Cobalt derivative {cy-[O-(Co-
PTPEG2)-O-CH2-]4}) by means of MALDI-TOF mass
spectrometry, NMR, UV–vis spectroscopy, and circular dichro-
ism, is here reported. Preliminary data about the formation of
complexes with Gramicidin S (a polypeptide having a relevant
pharmacological importance) are also discussed.
Synthesis of 5,15-Di-[p(x-methoxypolyethyleneoxy)phenyl]-
10,20-[p-hydroxyphenyl] Porphyrin, HO(H2-PTPEG2)OH
HO(H2-PTPEG2)OH was obtained by reaction between x-
methoxypolyethyleneoxy chloride [PEGMEC, having an aver-
age Mw ¼ 750 Da (MWD ¼ 1.05) and formed by reaction
between poly(ethyleneglycol)methyl ether (PEGME) and thi-
onyl chloride in THF] and H2-P(OH)4.21
Briefly, 2.625 g of PEGMEC (3.5 mmol) dissolved in 10 mL of
a H2O/THF (1/1) mixture, 0.298 g of H2-P(OH)4 (0.44
mmol) dissolved in 35 mL of a 0.5M NaOH aqueous solution
was added and the mixture refluxed for 24 h. Further 10 mL
of 0.5M NaOH and 10 mL of THF were then added and the
solution refluxed for other 24 h.
EXPERIMENTAL
Materials
When, examining aliquots of the mixture, the conversion in
di-(p-hydroxyphenyl)porphyrin, considering the relative
amount of the cluster of molecular ion peaks centered at
about m/z 2200 in the MALDI-TOF spectrum (see Fig. 1)
with respect to the clusters of other derivatives21 was judged
optimal, the solution was slightly acidified with CH3COOH,
dried under vacuum and the residue dissolved in CH2Cl2, fil-
tered and fractionated by column chromatography by using
silica gel as stationary phase and a solution of CH2Cl2/EtOH/
N(C2H5)3 (96.5/2.0/1.5) as an eluent. By MALDI-TOF-MS
(Fig. 2), 1H-NMR, COSY, and ROESY experiments, the third
All the solvents and basic materials were commercial prod-
ucts appropriately purified before use.
UV-Visible and Circular Dichroism
UV–visible spectra were recorded at room temperature by a
Shimadzu Model 1601 spectrophotometer, in quartz cells,
using tetrahydrofurane (THF) or H2O as a solvent. Circular
dichroism measurements were performed on a Jasco J815
spectropolarimeter at T ¼ 25.0 6 0.1ꢀC.
1H-NMR Analyses
1H-NMR, COSY, and T-ROESY spectra were obtained on a
UNI-
TYINOVA Varian instrument operating at 500 MHz (1H) using
VNMR for software acquisition and processing. Samples were
dissolved in CD2Cl2 and the chemical shifts expressed in
ppm by comparison with the CH2Cl2 residue signal. The
spectra were acquired at 27ꢀC, with a spin lock time of 0.5 s.
COSY and T-ROESY spectra were acquired using a Varian
standard impulse sequence. In the T-ROESY experiment, a
spin lock field of 2000 Hz with a spin lock time of 0.5 s
were applied.
MALDI-TOF Mass Spectrometric Analysis
Positive MALDI-TOF mass spectra were acquired by
a
Voyager DE-STR (PerSeptive Biosystem) using a simultane-
ous delay extraction procedure (25 kV applied after 2600 ns
with a potential gradient of 454 V mmꢁ1 and a wire voltage
of 25 V) and detection in linear mode.15,16
The instrument was equipped with a nitrogen laser (emis-
sion at 337 nm for 3 ns) and a flash AD converter (time
base 2 ns). Trans-3-indoleacrylic acid (IAA) was used as a
FIGURE 1 MALDI-TOF spectra of the reaction product between
PEGMEC and tetrakis-(p-hydroxyphenyl) porphyrin.
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FIGURE 2 MALDI-TOF mass spectrum of HO(H2-PTPEG2)OH.
compound eluted from the column (collected with a yield of
13% with respect to the initial porphyrin amount) resulted
pure HO(H2-PTPEG2)OH.
H, methylene protons c); a broad triplet at 3.89 ppm (4 H,
methylene protons d); some broad signals in the range 3.75–
3.55 ppm (128 H, PEO methylene protons); a singlet at 3.46
ppm (6 H, terminal methyl groups x); a singlet (not shown)
at ꢁ2.86 ppm (2H, NAH pyrrole protons 21, 22).
Its 1H-NMR spectrum (500 MHz, in CD2Cl2 at 27ꢀC; for the
atom identification see inset of Fig. 3) showed the following
signals: two unresolved doublets at 8.87 ppm (4 H, pyrrole
protons 2, 8, 12, 18) and 8.70 ppm (4 H, pyrrole protons 3,
7, 13, 17); a doublet at 8.01 ppm (4 H, phenyl protons a0); a
broad signal at 7.8 ppm (4 H, phenyl protons a); a doublet
at 7.27 (4 H, CH phenyl protons b0); a broad signal at 6.96
(4 H, CH phenyl protons b); a broad triplet at 4.12 ppm (4
Instead, the fourth compound eluted from the column
(collected with a yield of 28% with respect to the initial por-
phyrin amount) resulted the 5,10-di-[p(x-methoxypolyethy-
leneoxy)phenyl]-15,20-[p-hydroxyphenyl]porphyrin, HO(H2-
PCPEG2)OH (see Supporting Information for its 1H-NMR
characterization data).
FIGURE 3 1H-NMR of HO(H2-PTPEG2)OH. In the inset, structure of HO(H2-PTPEG2)OH and NMR assignments.
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SCHEME 1 Synthesis of tetra{5,15-di-[p(x-methoxypolyethyleneoxy)phenyl]-10,20-[p-oxyphenyl] methylen porphyrin}, cy-[O-(H2-
PTPEG2)-O-CH2-]4.
Synthesis of Cyclic Tetra{5,15-di-[p(x-
silica gel as stationary phase and a CHCl3/C2H5OH/N(C2H5)3
(96,5/2,0/1,5) mixture as eluent (yield of almost 100%).
methoxypolyethyleneoxy)phenyl]-10,20-[p-oxyphenyl]
Methylen Porphyrin}, cy-[O-(H2-PTPEG2)-O-CH2-]4
Cyclic-ethers, having methylene bridges between the por-
phyrin units, were synthesized by interfacial etherification
reaction in toluene/H2O between HO(H2-PTPEG2)OH
and a large excess of dibromomethane with tetrabutyl
ammoniumbromide (TBAB) used as phase-transfer agent
(Scheme 1).
RESULTS AND DISCUSSION
The synthesis of cyclic-ether oligomers containing porphyrin
units involved the use of HO(H2-PTPEG2)OH in turn
obtained by the reaction between H2-P(OH)4 and x-methyl-
oxy polyethyleneoxy chloride. As expected, from this last
reaction, a mixture of porphyrin derivatives with a different
number of polyethylenoxy branches was obtained.
In a typical synthetic procedure (see Scheme 1), to 200 mg
of HO(H2-PTPEG2)OH (ca. 0.095 mmol), dissolved in 100
mL of a 2M NaOH solution, 230 mg of TBAB (0.7 mmol) was
added at room temperature. Then, a solution of 10 mL of
CH2Br2 in 80 mL of toluene was added and the mixture
refluxed under vigorous stirring, monitoring the reaction by
TLC chromatography, and MALDI-TOF mass spectrometry. Af-
ter 24 h, the reaction was stopped and the material con-
tained in the organic phase recovered by rotoevaporation in
vacuum. From the mixtures of oligomers (yield of 90% with
respect to the initial HO(H2-PTPEG2)OH amount), the pure
cy-[O-(H2-PTPEG2)-O-CH2-]4 was collected by column chro-
matography, using silica gel as stationary phase and a
CHCl3/MeOH/N(C2H5)3 (97.5/2.0/0.5) mixture as eluant
(yield of 18%).
In fact, the MALDI-TOF spectrum of the obtained product
resulted constituted of five clusters of peaks (see Fig. 1): the
cluster centered at about m/z 800 was due to the unreacted
PEGMEC; the cluster centered at about m/z 1400 corre-
sponded to the oligomers of the mono-branched porphyrin
derivatives {the 5-[p-(x-methoxy-polyethylenoxy)phenyl]-
10,15,20-tri–(p-hydroxyphenyl) porphyrin}; the cluster cen-
tered at about m/z 2200 was due to the mixture of the two
di-branched porphyrin isomers {HO(H2-PTPEG2)OH and
the 5,10-di-[p-(x-methoxy-polyethylenoxy)phenyl]-15,20-di–
(p-hydroxyphenyl)porphyrin}; the cluster centered at about
m/z 2900 was due to the tri-branched porphyrin oligomers
{the 5,10,15-tri-[p-(x-methoxy-polyethylenoxy)phenyl]-20-(p-
hydroxyphenyl)porphyrin}; the last cluster centered at about
m/z 3600 was due to the tetra-branched porphyrin
derivative.
In particular, its 1H-NMR (500 MHz, in CD2Cl2 at 27ꢀC; for
the atom identification see Figure SI3 in Supporting Informa-
tion) showed: some broad signals in the range 9.10–8.67
ppm (32 H, CAH pyrrole protons) and 8.45–8.01 ppm (32 H,
phenyl protons); two broad signals at 7.70 and 7.34 ppm
(32 H, phenyl protons); a broad signal at 6.34 ppm (8 H,
methyleneoxy protons); some broad signals between 4.78
and 3.23 ppm (544 H, methylene protons of PEO); a singlet
at 3.18 ppm (24 H, methyloxy protons); a singlet at ꢁ2.71
ppm (8H, NAH pyrrole protons).
Pure HO(H2-PTPEG2)OH was obtained by chromatographic
fractionation and, on the basis of its MALDI-TOF mass spec-
trum and some marked characteristics of its NMR spectra,
HO(H2-PTPEG2)OH resulted the third eluted product. Figure
2 shows its MALDI-TOF mass spectrum consisting in a single
cluster of peaks (ca. 1600–2700 Da), centered at about m/z
2100 (ca. Mn ¼ 2090, MWD ¼ 1.04) with a separation of the
peaks of 44 amu as a consequence of the different total
number of oxyethylene units in the two PEG branches. In
particular, each molecular ion with a n þ m value between 0
and 25þ (see structure in the inset of Fig. 3) appears as
Preparation of the Cyclic tetra{5,15-di-
[p(x-methoxypolyethyleneoxy)phenyl]-10,20-
[p-oxyphenyl] Methylene-cobalt-porphyrin},
cy-[O-(Co-PTPEG2)-O-CH2-]4
The insertion of cobalt ions, by substitution of the two
hydrogen atoms in each porphyrin core of Cy-[O-(H2-
PTPEG2)-O-CH2-]4 was obtained by reaction of this last with
a large molar excess (1:10) of cobalt acetate in pyridine at
100ꢀC under a N2 atmosphere. Pure cy-[O-(Co-PTPEG2)-O-
CH2-]4 was recovered by chromatographic separation using
M
M
nþmH (peaks at m/z 1587 þ n44, indicated as ‘‘þ’’),
nþmNaþ (peaks at m/z 1609þ n44, indicated as ‘‘#’’) and
MnþmKþ (peaks at m/z 1625þ n44, indicated as ‘‘*’’) species.
The exact identification of HO(H2-PTPEG2)OH as the center
-symmetrical 10,20-di[p-hydroxyphenyl]porphyrin isomer,
was achieved by the NMR analysis (see Experimental section
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FIGURE 4 MALDI-TOF mass spectrum of oligo[5,15-di-[p(x-
methoxy-polyethyleneoxy)phenyl]-10,20-[p-oxyphenyl]
meth-
FIGURE 5 MALDI-TOF mass spectrum of cycle(tetra[5,15-di-
[p(x-methoxy-polyethyleneoxy) phenyl]-10,20-[p-hydroxyphenyl]
methylen porphyrin]).
ylen porphyrin].
protons with four Coþþ was confirmed by the shift of about
230 amu of the peak signals observed in its MALDI-TOF
spectrum (Fig. 6) with respect the signals of Figure 5.
and Fig. 3). The structural assignment was also supported by
T-ROESY and COSY experiments (see Supporting Information)
which showed, respectively, the diagnostic cross-peak
correlations between protons c and b; a and (3, 7)-(13, 17)
and a0 and (2,18)-(8, 12) and between the signals of the a-b,
a0-b0, and c-d protons.
The UV–vis spectra of cy-[O-(H2-PTPEG2)-O-CH2-]4 in THF
(continuous line) and in water (continuous line in the inset)
solutions are shown in Figure 7; they exhibited the character-
istic intense Soret-band (kmax ¼ 425 nm in THF and 424 nm
in water) and four satellite Q-bands (between 500 and 700
nm). It particular, the spectrum in THF resulted very similar
to that of the starting HO(H2-PTPEG2)OH (Fig. 7, dotted line;
kmax ¼ 421 nm in THF). Instead, the Soret band profile of
HO(H2-PTPEG2)OH in aqueous solution (see inset Fig. 7, dot-
ted line) clearly shows the energy shifts of the nearly degen-
erate monomer band, originated from the formation of H-type
(band at ca. 400 nm) and, mainly, J-type (band at ca. 442 nm)
aggregates.22,23 This phenomenon, as discussed in our recent
works,22,23 is due to the balance between the hydrophobic
interactions and the hydrogen bonding contribution that
determining the formation of J- or H-type aggregates.
To enhance the formation of cyclic oligomers, the subsequent
condensation reaction between HO(H2-PTPEG2)OH and
dibromomethane, was performed in diluted condition (for
more details see Experimental section).
Figure 4 shows the MALDI-TOF mass spectrum of the
oligomers mixture, which consists of several clusters of
peaks corresponding to the molecular ions of cyclic cy-[O-
(H2-PTPEG2)-O-CH2-]n oligomers with a different number of
repetitive units (p ¼ 3–5, see Scheme 1). As expected, the
higher intensity of the signals of the cluster relative to the
cyclic tetra-ether oligomer (centered at ca. 8400), indicated
this as the most abundant species formed in the reaction.
Pure cy-[O-(H2-PTPEG2)-O-CH2-]4 was then obtained by chro-
matographic column fractionation with a yield of 18%. Its
MALDI-TOF spectrum, reported in Figures 5, showed peaks
at m/z: 7537 þ n44, between about m/z 7500 to m/z 9200,
with n ¼ 0–36, corresponding to molecular ions detected
as MHþ.
The chemical structure of cy-[O-(H2-PTPEG2)-O-CH2-]4 was
also confirmed by NMR analysis (see Supporting Informa-
tion). In particular, the signal at 6.34 ppm was indicative of
the ether linkage formation14 between the porphyrins. More-
over, the spectrum of the cyclic tetramer showed, with
respect to that of the HO(H2-PTPEG2)OH, a broadening of
the NMR signals due to both the oligomeric structure of the
tetramer and the different conformations of the macro-cycle.
The synthetic procedure ended with the transformation of a
part of cy-[O-(H2-PTPEG2)-O-CH2-]4 in the corresponding
cobalt derivative (cy-[O-(Co-PTPEG2)-O-CH2-]4) by reaction
with cobalt acetate. The substitution of the eight pyrrole
FIGURE 6 MALDI-TOF spectrum of cy-[O-(Co-PTPEG2)-O-CH2-]4.
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FIGURE 7 UV–vis spectra of solutions in THF of cy-[O-(H2-
PTPEG2)-O-CH2-]4 (continuous line), HO(H2-PTPEG2)OH (dotted
line), and cy-[O-(Co-PTPEG2)-O-CH2-]4 (dashed line). In the
inset, UV–vis spectrum of aqueous solution of cy-[O-(H2-
PTPEG2)- O-CH2-]4 (continuous line) and HO(H2-PTPEG2)OH
(dotted line).
Differently, the UV–vis spectrum of the cy-[O-(Co-PTPEG2)-O-
CH2-]4 in THF solution (dashed line of Fig. 7) showed the
typical feature of a metalled porphyrin derivative, with rele-
vant changes in the position (a red shift) and intensity of the
Soret band and the disappearance of two Q-bands.
FIGURE 9 (a) ICD and (b) HT spectra of cy-[O-(Co-PTPEG2)-O-
CH2-]4 (concentration 5 ꢂꢃ10ꢁ7 M, dotted line) and its mixtures
with Gramicidin S (continuous line) in water solution (molar ra-
tio 1:100). In the inset, ICD spectra of Co-P/Gramicidin/Co-P
complex23 (concentration 6 ꢂꢃ10ꢁ6 M).
The decreased Q-band number was interpreted on the basis
of the already reported so-called ‘‘four-orbital model’’24 that
describes the low-lying (p, p*) excited states of porphyrins
in terms of electronic transitions between the two topmost
filled molecular orbitals (HOMO’s), a2u (p) and a1u (p), to
two degenerate lowest empty molecular orbitals (LUMO’s),
eg (p*). According to this model, metallo-porphyrins should
show only two visible Q-bands instead of the four Q-bands
shown by the free-porphyrin base.
To test the ability of cy-[O-(H2-PTPEG2)-O-CH2-]4 and cy-[O-
(Co-PTPEG2)-O-CH2-]4 to entrap polypeptide compounds,
experiments with Gramicidin S were performed. Besides its
well-known antibiotic properties, the choice fell on this cyclic
decapeptide [constituted of two identical series of five AAs:
(Val, Orn, Leu, D-Phe, and Pro)2] for its molecular size suita-
ble for its possible accommodation into the cyclic-ether cav-
ity (see inset of Fig. 8).
As expected, considering the marked ability of cobalt-por-
phyrin derivatives to complex AAs residue,21,25,26 significant
results were obtained with cy-[O-(Co-PTPEG2)-O-CH2-]4.
The Soret regions of the high tension voltage (HT, which is
roughly proportional to absorbance) and of the CD spectra of
freshly prepared aqueous solutions (5 ꢂ 10ꢁ7 M) of pure cy-
[O-(Co-PTPEG2)-O-CH2-]4 (dashed line) and its mixture with
Gramicidin S (1:100) (continuous line) are compared in
Figure 9. Differently from the trace in organic solvent (dashed
line in Fig. 6) the Soret band of cy-[O-(Co-PTPEG2)-O-CH2-]4
in water solution [dashed line in Fig. 9(b)] consisted of two
overlapped signals of comparable intensity with kmax values,
respectively, at about 415 and 430 nm. As previously
explained in similar cases,21 these absorptions can be
ascribed to the equilibrium between free cy-[O-(Co-PTPEG2)-
FIGURE 8 Sketch 3D of the host/guest complex. In the inset,
chemical structure of Gramicidin S.
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O-CH2-]4 molecules and their complex species with a different
number of ligands (H2O, AOH, etc.) bound to the metal atoms
in axial positions with respect to the porphyrin planes.
the ICD signal (Fig. 9) showed a partial asymmetric profile
because the appositive box’s walls, for the quadrate geome-
try, are at wide distance (ca. 21.2 Å) to allow an efficacy
excitonic coupling.27 Moreover, also the mutual arrangement
of the adjacent perpendicular porphyrins do not allow an
efficacy excitonic coupling.27,28
The presence of Gramicidin S caused a slight shift of kmax of
the two band [continuous line spectrum in Fig. 9(b)], respec-
tively, at 420 and 438 nm, which appeared also with a differ-
ent intensity.
CONCLUSIONS
Because cy-[O-(Co-PTPEG2)-O-CH2-]4 is an achiral compound,
as expected, no signal was observed in its CD spectrum
[dashed line in Fig. 9(a)] whereas an induced circular dichro-
ism (ICD) trace, consisting of an asymmetrical bisigned sig-
nal appearing in correspondence with the Soret absorption
band, was observed in its aqueous solution with Gramicidin
S [molar ratio 1:100; continuous curve of Fig. 9(a)]. The
symmetry breaking of the molecular species was considered
a strong confirmation of the formation of stable complexes
In summary, we here report the synthesis and the chemical
and spectroscopic characterization of a novel water soluble
cyclic tetra-ether porphyrin, cy-[O-(H2-PTPEG2)-O-CH2-]4,
having a molecular architecture similar to a nanobox, and of
its Cobalt derivative cy-[O-(Co-PTPEG2)-O-CH2-]4. Remark-
ably, CD experiments on a mixture in water solution of this
last and Gramicidin S showed the formation of supramolecu-
lar structures.
Really, the ICD signal was little intense, probably for the low
amount of cy-[O-(Co-PTPEG2)-O-CH2-]4/Gramicidin S com-
plex species formed, but sufficient enough to observe the
phenomenon. However, in this respect, a future goal will be
an increment of the amount of guest/host species modifying
the experimental conditions. It will also be need to verify the
true possible use of cy-[O-(Co-PTPEG2)-O-CH2-]4 as a drug
delivery system. In particular, considering the strong affinity
of porphyrins toward tumoral tissues, experiments about the
host/guest formation with anti-cancer drugs and its succes-
sive release will be proved.
between cy-[O-(Co-PTPEG2)-O-CH2-]4 and Gramicidin
molecules.
S
Furthermore, by comparison between CD and absorption
spectra, it was also possible assigned the HT signal centered
at 415 nm in the spectrum of pure cy-[O-(Co-PTPEG2)-O-
CH2-]4 [dashed line in Fig. 9(b)], to the free molecules
because no ICD signal was observed in its correspondence.
Otherwise, the two different ICD signals centered at 428 nm
(negative signal) and 448 nm (positive signal) were consid-
ered, as in previous similar cases,21,25,26 indicative of two
diverse cy-[O-(Co-PTPEG2)-O-CH2-]4/GramicidinS complexes;
probably, by the overlap of the corresponding ICD signals
could originate the asymmetry of the bands observed in
Figure 9(a).
ACKNOWLEDGMENTS
ꢀ
The authors thank the Ministero Istruzione Universita e Ricerca
(MIUR, Roma) for the partial financial support (PRIN 2008—
‘‘New Porphyrins as Chirogenetic Probes to Recognition of Pep-
tides and Proteins of Biological Interest’’—N. Prot.
2008KHW8K4) and by the project PON01_00074-DIATEME.
To put in evidence the effect of the spatial geometry of the
macromolecular box in the interaction with Gramicidin S, a
new experiment was performed using a mono-porphyrin sys-
tem, the 5,10,15,20-tetrakis-p-(x-methoxypolyethyleneoxy-
phenyl)-Cobalt-porphyrin23 (Co-P, a porphyrin with four PEG
branches bound in the peripheral position). The ICD spec-
trum of the Co-P/Gramicidin solution, consisting in a strong
[DCD ca. 29 mdeg; where DCD ¼ CD453 – CD440] and almost
symmetrical bisigned signal, is reported in the inset of Fig-
ure 9. It can be considered that among the AAs of the Grami-
cidin only Phe (by p–p interactions) and Orn (by interaction
between its d-amino group and the Cobalt atom) could be
able to interact with Co-P.
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