Synthesis, FT-IR, 1H, 13C NMR, ESI MS and PM5 studies of a new Mannich base of polyether antibiotic-Lasalocid acid and its complexes with Li+, Na+ and K+ cations

Article abstract of DOI:10.1016/j.saa.2012.11.106

The polyether antibiotic Lasalocid acid has been converted to its Mannich base derivative by a chemoselective one-pot reaction with formaldehyde and morpholine through the decarboxylation process. Spectroscopic studies of the structure of this new derivative have shown that in this ortho-phenol Mannich base the OH...N intarmolecular hydrogen bond is present. The compound forms complexes with Li⁺, Na⁺ and K⁺ cations of exclusively 1:1 stoichiometry. The structures of these complexes have been studied and visualized by semi-empirical calculation based on results of spectrometric and spectroscopic investigation. It is demonstrated that in contrast to Lasalocid acid the novel Mannich type derivative forms preferential complexes with Li⁺ cation.

Full text of DOI:10.1016/j.saa.2012.11.106

Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 104 (2013) 497–504
Contents lists available at SciVerse ScienceDirect
Spectrochimica Acta Part A: Molecular and
Biomolecular Spectroscopy
journal homepage: www.elsevier.com/locate/saa
1
Synthesis, FT-IR, H, 13C NMR, ESI MS and PM5 studies of a new Mannich base
of polyether antibiotic – Lasalocid acid and its complexes with Li+, Na+ and K+
cations
a,
a
a
b
⇑
´
Adam Huczynski , Jacek Rutkowski , Bogumil Brzezinski , Franz Bartl
a Faculty of Chemistry, Adam Mickiewicz University, ul. Umultowska 89b, 61-614 Poznan´, Poland
b Institute of Medical Physics and Biophysics, Charité, Universitätsmedizin Berlin, Charitéplatz 1, 10117 Berlin, Germany
g r a p h i c a l a b s t r a c t
h i g h l i g h t s
"
"
"
"
A new ortho-phenol Mannich base of
Lasalocid was synthesized by
chemoselective one-pot reaction.
The compound obtained is a useful
ligand for complexation of
monovalent cations.
Spectroscopic characterization of the
ligand and prepared complexes is
given.
O
O
O
O
OH
N
N
O
H
M+
O
N
HO
HO
H
H
H
OH O
O
O
OH O
O
OH O
M+
O
O
O
2-Li+, 2-Na+, 2-K+
Mannich derivative
Lasalocid acid (1)
O
OH
OH
H
2
of Lasalocid ( )
complexes
Semiempirical calculations of
structures of the complexes studied
are presented.
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 20 August 2012
Received in revised form 24 November 2012
Accepted 29 November 2012
Available online 8 December 2012
The polyether antibiotic Lasalocid acid has been converted to its Mannich base derivative by a chemose-
lective one-pot reaction with formaldehyde and morpholine through the decarboxylation process. Spec-
troscopic studies of the structure of this new derivative have shown that in this ortho-phenol Mannich
base the OAHꢀ ꢀ ꢀN intarmolecular hydrogen bond is present. The compound forms complexes with Li+,
Na+ and K+ cations of exclusively 1:1 stoichiometry. The structures of these complexes have been studied
and visualized by semi-empirical calculation based on results of spectrometric and spectroscopic inves-
tigation. It is demonstrated that in contrast to Lasalocid acid the novel Mannich type derivative forms
preferential complexes with Li+ cation.
Keywords:
Mannich base synthesis
Complexes with monovalent cations
Structural studies
Ó 2012 Elsevier B.V. All rights reserved.
Spectroscopic studies
Hydrogen bonds
Semiempirical calculations
Introduction
complexes with monovalent and divalent cations and transport
them across lipid bilayer. The influx of Na+ into the cell of Gram-
Lasalocid acid (Scheme 1) and its derivatives represent a large
class of ionophore antibiotics. These compounds show a broad
spectrum of bioactivity e.g. antibacterial, antifungal, antiparasitic
and antiviral [1–7]. Lasalocid acid sodium salt is used as an antibi-
otic for poultry and as a growth promoter for ruminants [1,2]. Lasa-
locid acid isolated from Streptomyces lasaliensis is able to form
positive bacteria leads to changes in pH and to an increase in the
osmotic pressure inside the cell, causing swelling and vacuoliza-
tion, eventually leading to cell death. The effectiveness of this pro-
cess strongly depends on the structure of the Lasalocid metal
cation complexes [1–4]. In previous studies we have shown that
the complex of Lasalocid with allylamine has higher anti-bacterial
activity than pure Lasalocid acid [8]. We found also that Lasalocid
acid and its complexes are strong cytotoxic agents towards cancer
cell lines. The cytostatic activity of Lasalocid and its complexes
⇑
Corresponding author.
E-mail address: adhucz@amu.edu.pl (A. Huczyn´ ski).
1386-1425/$ - see front matter Ó 2012 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.saa.2012.11.106

´
498
A. Huczynski et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 104 (2013) 497–504
Scheme 1. Mannich reaction conditions.
with amines against human cancer cell lines is higher than that of
cisplatin, indicating that Lasalocid and its complexes are promising
candidates for new anticancer drugs [9].
Synthesis of Mannich base of Lasalocid acid (2)
A mixture of Lasalocid (1 g, 1.69 mmol), paraformaldehyde
(253 mg, 8.45 mmol) and morpholine (736 mg, 8.45 mmol) in tol-
uene (200 mL) was stirred and heated under reflux for 5 h. Water
was collected in a Dean–Stark distilling trap during the reflux per-
iod. The resulting solution was diluted with petroleum ether and
transferred to the separatory funnel and washed once with water
and then one with 0.05 M HCl. The organic layer was evaporated
under reduced pressure giving 1.6 g of pale yellow resin, which
was then purified by dry column flash chromatography, giving
817 mg (75% yield) of final product. Elemental analysis calc. for
Since various N-functionalized morpholines show pharmaco-
logical activity, we synthesized a new morpholine Mannich base
derivative of Lasalocid acid. Such compounds are reported to exert
a number of important physiological activities such as antiemetic
or growth stimulant. They are also used in the treatment of inflam-
matory diseases, pain, migraine, and asthma [10–12]. In this con-
tribution, the nature of complexes formed between Mannich base
of Lasalocid acid with morpholine (2) and monovalent cations
(Li+, Na+ and K+) is studied using 1H NMR, 13C NMR, FTIR, ESI-MS
as well as PM5 semiempirical methods. The structures of these
complexes with metal cations are discussed in detail.
C38H63NO7: C 70.66%, H 9.83%, N 2.17%, O 17.34% found: C
70.64%, H 9.85%, N 2.14%, O 17.37%. The exemplary spectra are in-
cluded in the Supplementary material.
Experimental
Synthesis of 1:1 complexes of 2 with monovalent cations
Materials and methods
The 0.07 mol/L solutions of 1:1 complexes of 2 with monovalent
cations (Li+, Na+ and K+) were obtained by adding equimolar
amounts of MClO4 salt (M = Li, Na, K) dissolved in water-free ace-
Lasalocid sodium salt was isolated from veterinary premix –
AvatecÒ 20 (Alpharma Inc.), which contains about 20% pure Lasalo-
cid sodium salt. Formaldehyde, morpholine and the perchlorates
LiClO4, NaClO4 and KClO4 were commercial products of Sigma
and used without any further purification. Since the salts were hy-
drates, it was necessary to dehydrate them at several (6–10 times)
evaporation steps from a 1:5 mixture of acetonitrile and absolute
ethanol. The dehydration of perchlorates was followed by record-
ing of their FT-IR spectra in acetonitrile.
tonitrile (3.5 mL, c = 0.05 mol/L) to acetonitrile solution of
2
(3.5 mL, c = 0.05 mol/L). The solvent was evaporated under reduced
pressure to dryness and the oily residue was dissolved to the
appropriate volume (2.5 mL) using dry CH3CN or CD3CN,
respectively.
NMR measurements
CD3CN and CH3CN spectral-grade solvents were stored over 3 Å
molecular sieves for several days. Handling of the compounds was
performed in a carefully dried, CO2-free glove box.
The 1H and 13C NMR spectra of 2 and its complexes (0.07 mol/L)
with Li+, Na+ and K+ were recorded in CD3CN solutions using Bruker
Avance 600 MHz spectrometer. All spectra were locked to deute-
rium resonance of CD3CN. The 1H NMR measurements were carried
out at the operating frequency 600.0018 MHz and the 13C NMR
spectra at the operating frequency 150.885 MHz. The temperature
298.0 K and TMS as the internal standard were used in both cases.
No window function or zero filling was used. The errors of the 1H
and 13C NMR chemical shift values were 0.01 ppm and 0.1 ppm,
respectively. The 1H and 13C NMR signals were assigned using 2-
D (COSY, HETCOR, NOESY and HMBC) whose examples are shown
in the Supplementary materials (Figs. S1–S10). 2-D spectra were
recorded using standard pulse sequences from Bruker pulse-se-
quence libraries.
Isolation of Lasalocid sodium salt (1)
Lasalocid sodium salt was isolated from AvatecÒ 20 an anticoc-
cidial feed additive distributed by Alpharma Inc. 100 g of Avatec
was dissolved in dichloromethane. The solvent was evaporated un-
der reduced pressure and the crude product obtained was purified
by dry column vacuum chromatography (gradient solvent mixture
hexane/dichloromethane) giving 11 g pure Lasalocid sodium salt.
Preparation of Lasalocid acid
Lasalocid sodium salt (1.5 g) was dissolved in dichloromethane
(100 mL) and stirred vigorously with a layer of aqueous sulphuric
acid (100 mL, c = 0.75 mol/L). The organic layer containing 1 was
washed with distilled water and the dichloromethane was evapo-
rated under reduced pressure to dryness to produce the acid.
FT-IR measurements
The FT-IR spectra of 2 and its complexes with monovalent cat-
ions were recorded in acetonitrile solution (0.07 mol/L). A cell with

´
499
A. Huczynski et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 104 (2013) 497–504
Si windows and wedge-shaped layers was used to avoid interfer-
ences (mean layer thickness 170 m). The spectra were taken with
an IFS 113v FT-IR spectrophotometer (Bruker, Karlsruhe) equipped
with a DTGS detector; resolution 2 cmꢁ1, NSS = 64. The Happ-Gen-
zel apodization function was used.
and carboxylic group. Previous studies have shown that aminome-
thylation of phenols is achieved with formaldehyde and amines
under acidic conditions using the Mannich reaction, which occurs
readily in ortho- and para-positions affording substituted phenols
[20,21]. It has also been shown that alkyl keto group readily reacts
under acidic conditions [22,23]. However, Lasalocid is very unsta-
ble in acidic environment and higher temperatures. For this reason
we chose mild conditions for the Mannich reaction using only
three substrates (Lasalocid acid, morpholine and paraformalde-
hyde 1:1:1) to perform a one-pot reaction in toluene solutions un-
der reflux. Mannich base of Lasalocid acid (2) was easily isolated in
pure form after purification by dry column vacuum chromatogra-
l
Mass spectrometry
The ESI (Electrospray Ionization) mass spectra were recorded on
a Waters/Micromass (Manchester, UK) ZQ mass spectrometer
equipped with a Harvard Apparatus syringe pump. All samples
were prepared in acetonitrile. The measurements were performed
for the two types of samples: the solutions of 2 (5 ꢂ 10ꢁ5 mol/L)
with: (a) each of the cations Li+, Na+ and K+ (2.5 ꢂ 10ꢁ4 mol/L) ta-
ken separately and (b) the cations Li+, Na+ and K+ (5 ꢂ 10ꢁ5/3 mol/
L) taken together. Samples were introduced directly into the ESI
phy yielding
a pure product with relatively low 12% yield
(Scheme 1). The low yield of the reaction was unsatisfactory there-
fore we performed this reaction with five fold excess of morpholine
and paraformaldehyde. The change in stoichiometry of the reaction
gave a yield of 75%. Note that the chemoselectivity of this reaction
did not change and that the second aminomethyl group was not
introduced to the Lasalocid acid molecule under these reaction
conditions.
source using a Harvard pump at the flow-rate of 20 l
l minꢁ1. The
ESI source potentials were: capillary 3 kV, lens 0.5 kV, and extrac-
tor 4 V. The standard ESI mass spectra were recorded at the cone
voltages (cv) was 10, 30, 50, 70, 90, 110 V. The source temperature
was 120 °C and the desolvation temperature was 300 °C. Nitrogen
was used as the nebulizing and desolvation gas at flow-rates of 100
and 300 dm3 hꢁ1, respectively. Positive ion mode was selected for
mass spectrometric experiments. Full scans were recorded in the
mass range m/z 200–1000, and the mass resolution was of 1 unit.
The ESI MS spectra of the 1:1 complex of 2 with Li+, Na+, and K+ re-
corded at various cone voltage values are shown in the Supplemen-
tary material (Figs. S11–S14).
Synthesis of complexes of 2 with metal cations
The solutions of 1:1 complexes of 2 with monovalent cations
(Li+, Na+ and K+) were obtained by adding equimolar amounts of
MClO4 salt (M = Li, Na, and K) dissolved in acetonitrile to acetoni-
trile solution of 2. The solvent was evaporated under reduced pres-
sure to dryness and the oily residue was dissolved to the
appropriate volume using dry CH3CN or CD3CN, respectively. The
complexes have no tendency to crystallize and therefore were
studied in the solution.
PM5 semi-empirical calculation
The structures of product 2 and its complexes were determined
using the ESI-MS, FT-IR, 1H NMR, 13C NMR and PM5 semiempirical
methods and are discussed in detail herein. The 1H- and 13C NMR
signals were assigned using two-dimensional spectra such as
COSY, HETCOR, NOESY, HMBC shown in the Supplementary
materials.
PM5 calculations were performed using the Win Mopac 2007
program at the semiempirical level (Cache Work System Pro Ver-
sion 7.5.085 – Fujitsu) [13,14]. PM5 quantum semiempirical meth-
od uses the Schrödinger equation to determine bond strengths,
atomic hybridizations, partial charges, and orbitals from the posi-
tions of the atoms and the net charge.
Spectroscopic studies of 2
Results and discussion
FT-IR spectroscopy
In Fig. 1 the FT-IR spectrum of Lasalocid acid (1) (dashed line) is
compared with that of its Mannich base (2) (solid line). In the spec-
Synthesis
trum of 2 (solid line), the bands assigned to the
m(OAH) and
Synthesis of Mannich base of Lasalocid acid (2)
m
(C@O) stretching vibrations of the carboxylic group, present in
Design of procedures to obtain semi-synthetic derivatives of
Lasalocid acid (1) (Scheme 1) is challenging because its structure
is stable neither in strongly acidic nor in strongly basic conditions.
Additionally, since the structure of Lasalocid comprises many dif-
ferent functional groups (i.e. carboxylic, ketone, aromatic, etheric
hydroxyl groups) it was important for the synthetic transformation
to be highly chemoselective. Taking into account the above it was
deduced that a Mannich reaction would be ideal for conversion of
carboxylic group of Lasalocid to dialkylaminomethyl group since
the reaction can be carried out under neutral conditions and is
compatible with all of the other functional groups of Lasalocid.
The Mannich reaction is a classic method for the preparation of
nitrogen-containing compounds and therefore a very important
carbon–carbon bond-forming reaction in organic synthesis [15–
17]. It has been successfully employed as a key step in syntheses
of natural products and in medicinal chemistry [17–19]. When
chemically different reactive sites, capable of reacting indepen-
dently with the aminomethylating agent, are present in the sub-
strate’s molecule, the selectivity of the reaction is substantially
determined by the relative reactivity of each reactive centre. In
the Lasalocid acid molecule three types of reactive sites are pres-
ent, i.e. alkyl keto group, positions 5 and 6 in salicylic acid moiety,
the spectrum of Lasalocid acid at 3200–2700 cmꢁ1 and
1652 cmꢁ1, vanish completely indicating the absence of carboxylic
group with the formation of the respective Mannich base 2. The
band assigned to the m(C@O) vibrations of the ketone group is pres-
ent at 1712 cmꢁ1 in FT-IR spectra of 1 and 2 indicating that the
Mannich reaction was chemoselective and no transformation of
ketone group occurs.
1H and 13C NMR spectroscopy
In the 13C NMR spectrum of Mannich base 2 (Table S1), the most
characteristic signal of C(1) atom of the methylene group at N atom
was observed at 57.6 ppm, while the signal of C(1) atom of car-
boxyl group of Lasalocid acid (1) was at 173.2 ppm. The assign-
ment of the salicylic aromatic ring moiety was carried out on the
basis of two- and three-bond long-range correlation detected in
the HMBC spectrum. The correlation of the proton of methylene
group C(1)H2 at 3.76 ppm with the 13C NMR signals at 118.9 ppm
(C-2), 157.4 ppm (C-3) and 140.5 ppm (C-7) indicated the presence
of a morpholinemethyl group on C-2 atom (Table S1, Supplemen-
tary materials) and led to a conclusion that the morpholinemethyl
group is present at C-2 atom (Scheme 1).

´
A. Huczynski et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 104 (2013) 497–504
500
taining a morpholine moiety group due to the charge-remote frag-
100
50
0
mentation mechanism [31,32] giving the respective cation
observed at m/z 565, 581, 597, respectively. The ESI MS data of
the complexes of 2 indicate that no abstraction of water molecule
in the fragmentation process occurs, while dehydration process has
always been observed during the ESI MS fragmentation process of
Lasalocid and its ester derivatives. Furthermore, the ESI spectra of
the Li+, Na+ and K+ complexes with 2 show characteristic signals at
m/z 361, 377, 393, respectively, assigned to fragment ion C, formed
directly from the molecular cation, by cleavage of the C(11)AC(12)
bond. Previously, Lopes et al. proposed a mechanism of the forma-
tion of fragment ion C for complexes of the Lasalocid salts with
monovalent cations [26]. In this mechanism a six-membered inter-
mediate facilitates a Ei type b-elimination leading to cation C
(Scheme 2) [26–28] This mechanism is promoted by the monova-
lent metal cation withdrawing electron density from the carbonyl
group due to the ion–dipole interaction [28] and is characteristic of
Lasalocid salts and Lasalocid ester complexes [28–31].
(a)
2
1
4000 3500 3000 2500 2000 1500 1000 500
100
(b)
50
The ESI-MS spectrum of the 1:1:1:3 mixture of LiClO4, NaClO4,
KClO4 and 2, measured at cv = 10 V (Fig. S14) shows three signals
at m/z 652, 668 and 684 characteristic of the 1:1 complexes of 2
with Li+, Na+ and K+ cations, respectively. The intensity of the signal
assigned to the 2ꢁLi+ complex is the highest, clearly indicating that
2 preferentially forms complexes with Li+ cations. The intensities of
the signals of 2ꢁNa+ and 2ꢁK+ complexes are comparable, but sig-
nificantly lower than that of 2ꢁLi+ complex, demonstrating that 2
shows much lower affinity to Na+ and K+ cations.
1712
1750
1652
0
1800
1700
1650
1600
-1
1550
1500
Wavenumber [cm
]
Fig. 1. The FT-IR spectra of: 1 (- - -) and 2 (––), recorded in CH3CN: (a) 4000–
400 cmꢁ1; (b) 1800–1500 cmꢁ1
.
1H and 13C NMR spectroscopy
Table 1
Heat of formation (kcal/mol) of 2 and its complexes with the cations calculated by
PM5 method.
The most important differences
D(ppm) between the respective
chemical shifts observed in the spectrum of 2 and the spectra of its
1:1 complexes with Li+, Na+ and K+ cations all in CD3CN are col-
lected in Supplementary materials (Table S1). The signals assigned
to the O(4)H and O(8)H protons in the 1H NMR spectrum of 2 arise
as a doublet at 2.74 ppm and a singlet at 2.99 ppm, respectively,
indicating that both protons are involved in only weak hydrogen
bonds (Fig. 2). In the 1H NMR spectra of 1:1 complexes formed be-
tween 2 and LiClO4, NaClO4, and KClO4, the most shifted signals are
found at ca. 11.35 ppm, assigned to O(3)H proton of the phenolic
group, demonstrating that in the complexes the intramolecular
O(3)HꢀꢀꢀN hydrogen bond is still present. In these spectra the sig-
nals of O(4)H and O(8)H protons are separate (Fig. 2) and shifted,
depending on the kind of the cation, towards lower ppm values
indicating that the hydrogen bonds in which these groups are in-
volved, are weaker than and different from those present in the
structure of 2.
The interactions between the oxygen atoms of the molecule of 2
with Li+ cation are clearly indicated in the 13C NMR spectra by the
shifts of C(13) carbon signal of the ketone group from 214.8 ppm
(uncomplexed 2) to 223.8 ppm (2ꢁLiClO4 complex) and the shifts
of the signals of C(18), C(19), C(22) and C(23) carbon atoms (Tables
S1 and S2), i.e. the atoms which are close the O atoms coordinating
Li+ cation. The same applies to the chemical shifts of the signals of
all carbon atoms close to the O atoms which coordinate Na+ and K+
cations within the 1:1 complexes, although all the chemical shift
changes are significantly lower in comparison with those observed
in the 13C NMR spectrum of 2ꢁLiClO4 complex. These results dem-
onstrate that the interactions of 2 with Na+ or K+ cations are clearly
weaker than those with Li+ cation (Table S1), which is also in agree-
ment with the PM5 semiempirical calculations discussed below.
Complex
HOF (kcal/mol)
D
HOF (kcal/mol)
2
ꢁ378.16
ꢁ254.91
–
2+Liþuncomplexed
ꢁ72.21
ꢁ33.96
ꢁ29.47
2+Liþcomplexed
2+Naþuncomplexed
2+Naþcomplexed
2+Kþuncomplexed
2+Kþcomplexed
ꢁ327.12
ꢁ236.11
ꢁ270.07
ꢁ261.12
ꢁ290.68
D
HOF = HOF2+M
– HOF2+M uncomplexed, M-metal cation.
complexed
Phenolic Mannich bases having the aminomethyl group in the
ortho position form the OAHꢀꢀꢀN intramolecular hydrogen bond.
The equilibrium between hydrogen-bonded and proton transfer
forms was also observed and discussed by several authors [24,25].
In the 1H NMR spectrum of 2 the most significantly shifted sig-
nal at 11.20 ppm is assigned to the proton of the O(3)H group at
the aromatic ring involved in the intramolecular O(3)HꢀꢀꢀN hydro-
gen bond.
Spectroscopic studies of 1:1 complexes of 2 with Li+, Na+ and K+ cations
ESI mass spectrometry
The ESI MS spectra of the 1:1 complexes of 2 with Li+, Na+, and
K+ recorded at various cone voltage values are shown in the Sup-
plementary material in Figs. S11–S13 (Supplementary informa-
tion), respectively. The ESI mass spectra of the complexes of 2
with Li+, Na+ and K+, recorded at cone voltage 10 V, show intense
peaks at m/z 652, 668, 684 respectively, indicating the formation
of complexes of exclusively 1:1 stoichiometry. With increasing
cone voltage, the intensity of the m/z signals of the 1:1 complexes
(ion A in Scheme 2) decreases and the intensity of two fragment
ion signals increases as shown in Figs. S11–S13. The fragmentary
cation B is formed by abstraction of the part of the molecule con-
FT-IR spectroscopy
In Fig. 3 the FT-IR spectra of 2 and its 1:1 complexes with Li+,
Na+ and K+ are compared. In the spectrum of 2 (solid line) the
broad band assigned to the OAH stretching vibrations arising in
the range 3700–3250 cmꢁ1 shows a complex structure following

´
501
A. Huczynski et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 104 (2013) 497–504
O
N
CH2
H
O
O
charge-remote
fragmentation
O
O
O
O
O
O
O
O
H
H
M
M
A
B
HO
HO
M+ = Li+ [C38H63LiNO7]+ m/z 652
M+ = Li+ [C34H54LiO6]+ m/z 565
M+ = Na+ [C38 63
H
NNaO7]+ m/z 668
KNO7]+ m/z 684
M+ = Na+ [C34 54
M+ = K+ [C34 54
H
NaO6]+ m/z 581
M+ = K+ [C38 63
H
+
H
KO67
]
m/z 597
-elimination
(Ei-type)
-elimination
(Ei-type)
O
H
O
O
O
O
M
M
O
HO
HO
C
M+ = Li+ [C21H38LiO4]+ m/z 361
M+ = Na+ [C21 38
H
NaO4]+ m/z 377
KO4]+ m/z 393
M+ = K+ [C21 38
H
Scheme 2. The proposed in source fragmentation pathways of 2 complexes with monovalent cations (M = Li+, Na+, and K+). The calculated formula of fragment ion and exact
m/z values of fragment ion added for clarity.
Fig. 2. Parts of the 1H NMR spectra of: (a) 2, (b) 2ꢁLiClO4 complex, (c) 2ꢁNaClO4 complex, (d) 2ꢁKClO4 complex. The corresponding proton assignments to the structures of 2
and its complexes are shown.

´
A. Huczynski et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 104 (2013) 497–504
502
and is engaged in the coordination process of the metal cations.
100
50
0
The interaction of C(13)@O oxygen atom with Li+ cation is the
(a)
strongest.
2
PM5 calculations
2-LiClO
4
4
2-NaClO
2-KClO
On the basis of the above-discussed, spectroscopic data of the
structure of 2 is calculated using the PM5 semiempirical method
(Tables 1 and 2). In this structure, shown in Fig. 4, the O(3)H phe-
nolic group is hydrogen-bonded to the nitrogen atom of morpho-
line moiety in the Mannich base fragment and the C(13)@O(5)
keto group is not involved in a hydrogen bond. The calculated
parameters of the intramolecular O(3)HꢀꢀꢀN hydrogen bond present
in 2 are similar to those calculated previously for ortho-dimethy-
laminomethylphenol by Koll et al. [32]. The molecule of 2 forms
pseudo-cyclic structure stabilised by O(4)HꢀꢀꢀO(6) and O(8)HꢀꢀꢀO(4)
intramolecular hydrogen bonds.
The calculated heat of formation (HOF) of the structure of 2
with O(3)HꢀꢀꢀN hydrogen bond is lower than that of its alternative
structure without this bond or with a proton transferred to the
nitrogen atom as shown in Scheme S1 (Supplementary materials),
indicating that this structure is more favourable than that of both
alternative types. This result is in agreement with the experimental
FT-IR and 1H NMR data. The data collected in Table 1 also show
that the formation of complexes of 2 with the cations is energeti-
cally favourable. The calculated structures of 2 and its 1:1 com-
plexes with various monovalent cations are shown in Fig. 4.
All these calculated complex structures are stabilised by intra-
molecular hydrogen bonds whose parameters depend on the coor-
dinating cations (Table 2). This result is in agreement with the
conclusion from the 1H NMR and FT-IR spectra discussed above
(see Figs. 2 and 3). The structures of the 2ꢁLi+ and 2ꢁNa+ com-
plexes are stabilized by four intramolecular hydrogen bonds such
4
4000 3500 3000 2500 2000 1500 1000 500
100
(b)
(c)
3456
3490
3530
50
0
3500
3474
3500
3700
3600
3400
3300
100
50
0
1694
1712
1725
1706
1709
1750
1700 1675
1650
-1
1625
1600
Wavenumber [cm
]
Fig. 3. FT-IR spectra of: (––) 2, (—) 2ꢁLiClO4, (ꢀꢀꢀ) 2ꢁNaClO4, and (––) 2ꢁKClO4
recorded in CD3CN in the ranges of: (a) 4000–400 cmꢁ1; (b)
stretching vibrations.
m(OH) and (c) m(C@O)
as
O(3)AHꢀ ꢀ ꢀN(1),
O(8)AHꢀ ꢀ ꢀO(4),
O(8)AHꢀ ꢀ ꢀO(7)
and
O(4)AHꢀ ꢀ ꢀO(6) which are shown in Fig. 4 and their geometric
parameters are collected in Table 3. Additionally the structures of
the 2ꢁLi+ and 2ꢁNa+ complexes are stabilized by four oxygen
atoms coordinating the respective cation. In contrast, the 2ꢁK+
complex is stabilized by three hydrogen bonds and five oxygen
atoms coordinating K+ cation. The interatomic distances between
the coordinated cations and the coordinating oxygen atoms to-
gether with their partial charges are summarized in Table 2. These
data demonstrate that with increasing ionic radii of the cations the
coordination spheres of these cations change. Different are also the
interatomic distances between the cations and the coordinating
oxygen atoms, as well as their partial charges.
from the involvement of O(4)H and O(8)H hydroxyl groups in
intramolecular hydrogen bonds of various strength. The structure
of this broad band changes with the complexation process and
with the type of metal cation in the complex. The same is true with
the band assigned to C@O stretching vibration of the ketone group.
In the spectra of the 1:1 complexes of 2 with Li+ (dashed line), Na+
(dotted line), K+ (dash-dotted line) the respective bands arise at
1694 cmꢁ1, 1706 cmꢁ1 and 1709 cmꢁ1, respectively. This demon-
strates that the carbonyl group ceases to be hydrogen bonded
Table 2
The interatomic distances (Å) and partial charges for O atoms of 2 coordinating metal cations in complexes structures calculated by PM5 method.
Complex with monovalent
cation
Monovalent cation partial
charge
Coordinating
atom
Coordinating atom partial
charge
Distance (Å) coordinating
atom ? cation
2ꢁLi+
2ꢁNa+
2ꢁK+
+0.492
+0.573
+0.581
O(4)
O(5)
O(6)
O(7)
ꢁ0.443
ꢁ0.355
ꢁ0.379
ꢁ0.368
ꢁ0.424
ꢁ0.364
ꢁ0.399
ꢁ0.389
ꢁ0.437
ꢁ0.360
ꢁ0.396
ꢁ0.375
ꢁ0.440
2.11
2.03
2.00
2.02
O(4)
O(5)
O(6)
O(7)
2.37
2.32
2.34
2.31
O(4)
O(5)
O(6)
O(7)
O(8)
2.84
2.80
2.82
2.75
2.83

´
503
A. Huczynski et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 104 (2013) 497–504
Fig. 4. The structures of: (a) 2 and its complexes, (b) 2ꢁLi+, (c) 2ꢁNa+, (d) 2ꢁK+ calculated by PM5 semiempirical method (WinMopac 2007).
the structure of 2 the oxygen atom of the C@O ketone group is al-
ways involved in coordination process of metal cation, especially
with Li+ because it is demonstrated that the strongest intramolec-
ular hydrogen bonds are formed within structure of the 2ꢁLi+ com-
plex. In the complexes of 2 with monovalent cations we observe
that the nitrogen atom of Mannich base, phenolic oxygen atom
as well as morpholine moiety play no role in the complexation
process.
Table 3
The lengths (Å) and angles (°) of the hydrogen bond for 2 complexes calculated by
PM5 method.
Compound
Atoms engaged in hydrogen bonds
Length (Å)
Angle (°)
2
O(3)AHꢀ ꢀ ꢀN(1)
O(4)AHꢀ ꢀ ꢀO(6)
O(8)AHꢀ ꢀ ꢀO(4)
O(3)AHꢀ ꢀ ꢀN(1)
O(8)AHꢀ ꢀ ꢀO(4)
O(8)AHꢀ ꢀ ꢀO(7)
O(4)AHꢀ ꢀ ꢀO(6)
O(3)AHꢀ ꢀ ꢀN(1)
O(8)AHꢀ ꢀ ꢀO(4)
O(8)AHꢀ ꢀ ꢀO(7)
O(4)AHꢀ ꢀ ꢀO(6)
O(3)AHꢀ ꢀ ꢀN(1)
O(8)AHꢀ ꢀ ꢀO(4)
O(4)AHꢀ ꢀ ꢀO(6)
2.70
2.73
2.85
147
113
161
2ꢁLi+
2ꢁNa+
2ꢁK+
2.71
2.95
2.72
2.53
148
165
115
111
Acknowledgements
Financial support from budget funds for science in years 2012–
2013 – Grant ’’Iuventus Plus’’ of the Polish Ministry of Science and
Higher Education-No. 0179/IP3/2011/71, is gratefully acknowl-
edged. The financial support of the DAAD (The German Academic
2.71
2.97
2.70
2.60
148
160
114
109
´
Exchange Service) is gratefully acknowledged by Adam Huczynski.
2.71
2.46
2.97
148
106
140
Appendix A. Supplementary material
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.saa.2012.11.106.
Conclusions
References
For the first time the Lasalocid Mannich base derivative (2) and
its 1:1 monovalent metal cation complexes have been obtained
and characterized by spectroscopic (FT-IR, 1H- and 13C NMR), mass
spectrometry (ESI-MS) and PM5 semiempirical methods. Lasalocid
acid (1) a well known polyether antibiotic has been converted to its
Mannich base derivative (2) by a simple one-pot reaction. Spectro-
scopic studies of the structure of 2 have shown that in this ortho-
phenol Mannich base the O–HꢀꢀꢀN intarmolecular hydrogen bond
is present. The ability to form complexes with Li+, Na+ and K+ cat-
ions by 2 has been studied in detail. It has been demonstrated that
2 preferentially forms a complex with Li+ cations. The electrospray
ionization mass spectra indicated the formation of stable com-
plexes of 1:1 stoichiometry. With increasing cone voltage values
the fragmentation of the respective complexes is detected and is
connected with the Ei type b-elimination, charge-remote fragmen-
tation mechanism and dehydration processes. The formation of the
complexes as well as the intramolecular hydrogen bonds stabiliz-
ing their structures is demonstrated by 1H and 13C NMR, FT-IR
spectra and PM5 semiempirical calculations. It is shown that in
[1] R. Martineau, C. Benchaar, H.V. Petit, H. Lapierre, D.R. Ouellet, D. Pellerin, R.
Berthiaumef, J. Dairy Sci. 90 (2007) 5714.
[2] M. Mitrovic, E.G. Schildknecht, Poultry Sci. 54 (1975) 750.
[3] J.W. Westley, Polyether Antibiotics Naturally Occurring Acid Ionophores, 1,
Marcel Dekker Inc., New York, 1982.
[4] B.C. Pressman, Antibiotics and their complexes, 1, Marcel Dekker Inc., New
York, 1985.
[5] A. Kart, A. Bilgili, Anim. J. Vet. Adv. 7 (2008) 748.
[6] A. Huczyn´ ski, A. Doman´ ska, I. Paluch, J. Stefan´ ska, B. Brzezinski, F. Bartl,
Tetrahedron Lett. 49 (2008) 5572.
[7] R. Pankiewicz, D. Remlein-Starosta, G. Schroeder, B. Brzezinski, J. Mol. Struct.
783 (2006) 136.
[8] A. Huczyn´ ski, J. Janczak, J. Rutkowski, D. Łowicki, A. Pietruczuk, J. Stefan´ ska, B.
Brzezinski, F. Bartl, J. Mol. Struct. 936 (2009) 92.
´
´
[9] A. Huczynski, J. Rutkowski, J. Wietrzyk, J. Stefanska, E. Maj, M. Ratajczak-Sitarz,
A. Katrusiak, B. Brzezinski, F. Bartl, J. Mol. Struct. 1032 (2013) 69.
[10] J.J. Hale, S.G. Mills, M. MacCross, C.P. Dorn, P.E. Finke, R.J. Budhu, R.A. Reamer,
W.P. Huskey, D. Luffer-Atlas, B.J. Dean, E.M. McGowan, W.P. Feeney, S.H.L.
Chiu, M.A. Cascieri, G.G. Chicchi, M.M. Kurtz, S. Sadowski, E. Ber, F.D. Tattersall,
N.M.J. Rupniak, A.R. Williams, W. Raycroft, R. Hargreaves, J.M. Metzger, D.E.
Maclntyre, J. Med. Chem. 43 (2000) 1234.
[11] P. Avramova, N. Danchev, R. Buyukliev, T. Bogoslovova, Arch. Pharm. 331
(1998) 342.

´
A. Huczynski et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 104 (2013) 497–504
504
[12] R.S. Varma, R. Prakash, M.M. Khan, A. Ali, Indian Drugs 23 (1986) 345.
[13] CAChe 5.04 UserGuide, Fujitsu, 2003.
[22] N. Holy, R. Fowler, E. Burnett, R. Lorenz, Tetrahedron 35 (1979) 613.
[23] K. Manabe, S. Kobayashi, Org. Lett. 1 (1999) 1965.
[14] P. Przybylski, A. Huczyn´ ski, B.J. Brzezinski, J. Mol. Struct. 826 (2007) 156.
[15] H.T. Cao, T. Roisnel, A. Valleix, R. Gree, Eur. J. Org. Chem. (2011) 3430.
[16] C.-H. Zhao, L. Liu, D. Wang, Y.-J. Chen, Eur. J. Org. Chem. (2006) 2977.
[17] A. Ting, S.E. Schaus, Eur. J. Org. Chem. (2007) 5797.
[18] T. Hirose, T. Sunazuka, D. Yamamoto, E. Kaji, S. Omura, Tetrahedron Lett. 46
(2006) 6761.
[19] E.J. Corey, R.D. Balansom, J. Am. Chem. Soc. 96 (1974) 6516.
[20] K. Matsumoto, K. Joho, S. Mimori, H. Iida, H. Hamana, A. Kakehi, Heterocycles
76 (2008) 106.
[24] M. Rospenk, L. Sobczyk, Magn. Res. Chem. 27 (1989) 445.
[25] M. Rospenk, J. Mol. Struct. 109 (1990) 221.
[26] C. Cheng, M.L. Gross, Mass Spectrom. Rev. 19 (2000) 398.
[27] M.L. Gross, Int. J. Mass Spectrom. 200 (2000) 611.
[28] N.P. Lopes, P.J. Gates, J.P.G. Wilkins, J. Staunton, Analyst 127 (2002) 1224.
[29] A. Huczyn´ ski, J. Rutkowski, B. Brzezinski, Struct. Chem. 22 (2011) 627.
´
[30] A. Huczynski, M. Ratajczak-Sitarz, A. Katrusiak, B. Brzezinski, J. Mol. Struct. 998
(2011) 206.
[31] G.J. Francis, M. Forbes, D.A. Volmer, D.K. Bohme, Analyst 130 (2005) 508.
[32] A. Koll, S.M. Melikova, A. Karpfen, P. Wolschann, J. Mol. Struct. 559 (2001) 127.
´
[21] K. Bujnowski, A. Adamczyk-Wozniak, L. Synoradzki, Arkivoc 13 (2008) 106.