Spectroscopic studies, crystal structures and antimicrobial activities of a new lasalocid 1-naphthylmethyl ester

Article abstract of DOI:10.1016/j.molstruc.2008.04.037

A new lasalocid 1-naphthylmethyl ester (NAFA) has been synthesised and studied by X-ray, ¹H NMR, ¹³C NMR, FT-IR, UV-vis, fluorescence as well as by PM5 semiempirical methods. The crystals of NAFA belong to the monoclinic system with the space group P2₁ with a = 13.4251(5) ?, b = 17.1064(7) ?, c = 18.5454(7) ?, β = 98.924(4)° and Z = 4. Two conformers of NAFA have been observed for two symmetry-independent molecules in different crystal environments. The molecular conformation of NAFA is partially stabilized by three intramolecular hydrogen bonds, in which the keto group is not involved. The FT-IR spectrum of NAFA in chloroform indicates that in this solvent the equilibrium between two structures of NAFA is realized. In one of the structures, the keto group is hydrogen bonded while in the other one this group is not involved in any hydrogen bond. The two structures of NAFA are discussed in detail. The new ester which has been additionally tested for its antimicrobial properties shows a certain activity against Gram-positive bacteria, however no activity against Gram-negative bacteria and Candida.

Full text of DOI:10.1016/j.molstruc.2008.04.037

Journal of Molecular Structure 891 (2008) 481–490
Contents lists available at ScienceDirect
Journal of Molecular Structure
journal homepage: www.elsevier.com/locate/molstruc
Spectroscopic studies, crystal structures and antimicrobial activities of a new
lasalocid 1-naphthylmethyl ester
a
a
a
a
b
´
´
Adam Huczynski , Izabela Paluch , Małgorzata Ratajczak-Sitarz , Andrzej Katrusiak , Joanna Stefanska ,
Bogumil Brzezinski a, , Franz Bartl
c
*
a Faculty of Chemistry, Adam Mickiewicz University, Grunwaldzka 6, 60-780 Poznan, Poland
b Medical University of Warsaw, Department of Pharmaceutical Microbiology, Oczki 3, 02-007 Warsaw, Poland
c Institute of Medical Physics and Biophysics, Charité – Universitätsmedizin Berlin Campus, Charité Mitte, Ziegelstr. 5/9, 10117 Berlin, Germany
a r t i c l e i n f o
a b s t r a c t
A new lasalocid 1-naphthylmethyl ester (NAFA) has been synthesised and studied by X-ray, 1H NMR, 13
C
Article history:
Received 26 March 2008
Accepted 19 April 2008
Available online 29 April 2008
NMR, FT-IR, UV–vis, fluorescence as well as by PM5 semiempirical methods. The crystals of NAFA belong
to the monoclinic system with the space group P21 with a = 13.4251(5) Å, b = 17.1064(7) Å,
c = 18.5454(7) Å, b = 98.924(4)° and Z = 4. Two conformers of NAFA have been observed for two symme-
try-independent molecules in different crystal environments. The molecular conformation of NAFA is
partially stabilized by three intramolecular hydrogen bonds, in which the keto group is not involved.
The FT-IR spectrum of NAFA in chloroform indicates that in this solvent the equilibrium between two
structures of NAFA is realized. In one of the structures, the keto group is hydrogen bonded while in
the other one this group is not involved in any hydrogen bond. The two structures of NAFA are discussed
in detail. The new ester which has been additionally tested for its antimicrobial properties shows a cer-
tain activity against Gram-positive bacteria, however no activity against Gram-negative bacteria and
Candida.
Keywords:
Ionophores
Lasalocid esters
X-ray
Spectroscopy
Antimicrobial activity
Ó 2008 Elsevier B.V. All rights reserved.
1. Introduction
in solution. The methods we used to characterise it are: X-ray dif-
fraction, FT-IR, 1H NMR, 13C NMR, UV–vis, fluorescence-spectros-
Ionophorous antibiotics belong to a group of highly bioactive
molecules, because they are able to transport monovalent and
bivalent metal cations across natural and artificial lipid mem-
branes [1–11]. Lasalocid (LAS) isolated from Streptomyces lasalien-
sis is a well-known representative of these compounds. It is used as
a growth-promoting agent and as a coccidiostat in beef cattle,
sheep, chickens and turkeys [12–17].
Up to now some esters of lasalocid, in particular those with
polyoxaalkyl chains or aromatic substituents have been synthes-
ised and their structures have been studied by different methods
[18–23]. We could show that with increasing length of the oxaalkyl
chains the oxygen atoms become more active in the complexation
process of the monovalent cations. For the esters with aromatic
substituents no influence of these rings on the complexation pro-
cess was observed. However, this kind of derivatization probably
increases the formation of crystal formation which can then be
studies by the X-ray method.
copy and PM5. The antimicrobial activity of this compound is
also investigated.
2. Experimental
2.1. Preparation lasalocid 1-naphthylmethyl ester (NAFA)
Lasalocid sodium salt was extracted from Avatec (Fa. Spezialf-
utter Neuruppin). Lasalocid sodium salt was dissolved in dichlo-
romethane and stirred vigorously with
a layer of diluted
aqueous sulphuric acid (pH 1.5). The lasalocid (LAS) containing
the organic layer was washed with distilled water. Subsequently,
dichloromethane was evaporated under reduced pressure to
dryness.
A
mixture of the 1-(chloromethyl) naphthalene (175 mg,
1.00 mmol, Fluka), LAS (500 mg, 0.85 mmol) and 1,8-diazabicy-
clo[5.4.0]undec-7-en (DBU) (130 mg, 0.85 mmol) and 40 ml tolu-
ene was heated at 90 °C for 5 h. After cooling, the precipitate
DBU–hydrochloride (DBUꢀHCl) was filtered and washed with hex-
ane. The filtrate was evaporated under reduced pressure and the
residue purified by chromatography on silica gel (Fluka type 60)
yielding NAFA (300 mg, 48% yield, mp 115 °C). The crystals of NAFA
were obtained by crystallization from dried acetonitrile solutions.
In the present contribution, we report on the structure of a new
lasalocid 1-naphthylmethyl ester (Scheme 1) in the solid state and
* Corresponding author. Tel.: +48 618291330.
E-mail address: bbrzez@main.amu.edu.pl (B. Brzezinski).
0022-2860/$ - see front matter Ó 2008 Elsevier B.V. All rights reserved.
doi:10.1016/j.molstruc.2008.04.037

´
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A. Huczynski et al. / Journal of Molecular Structure 891 (2008) 481–490
40
39
38
41
45
42
43
37
35
36
44
31
O(2)
O(1 )
8
1
32
12
O
O
33
30
29
2
16
HO
3
7
15
13
11
14
10
O(3 )
17
9
28
21
34
O
O(6)
OH
O(4)
O
O(5)
6
4
18
19
27
20
H3C
5
O(7 )
24
O
26
23
25
22
OH
O(8)
Scheme 1. The structures and atom numbering of NAFA.
Elemental analysis: Calculated: C, 73.94%; H, 8.55%. Found: C,
Table 1
Crystal data and structure refinement
73.88%; H, 8.67%.
Empirical formula
Formula weight
Temperature (K)
Wavelength
Crystal system, space group
Unit cell dimensions
a (Å)
C45H62O8
730.95
293(2)
0.71073 Å
Monoclinic, P21
2.2. X-ray measurements
The crystals selected for single-crystal X-ray diffraction mea-
surements formed colourless parallelepipeds with well-devel-
oped faces. They were stable under normal conditions and the
X-ray diffraction measurements were carried out on a Kuma
KM-4 CCD diffractometer at room temperature. The structure
was solved by direct methods [24] and refined by full-matrix
least squares [25]. The positions of all the H-atoms were deter-
mined on the basis of the molecular geometry (CAH 0.98–0.93,
OAH 0.82 Å) and their Uiso’s were related to the thermal vibra-
tions of their carriers. The absolute conformation of NAFA in
the crystal was consistent with the chemical information. Se-
lected details about the crystal structure, experiment and struc-
ture solution and refinement are given in Table 1 and the
fractional atomic coordinates are listed in Table 2. The crystallo-
graphic-information-file (CIF) has been deposited at the Cam-
13.4251(5)
17.1064(7)
18.5454(7)
98.924(4)
b (Å)
c (Å)
b (°)
Volume (Å3)
4207.5(3)
Z
4
Calculated density
Absorption coefficient
F(000)
1.154 g cmꢁ3
0.078 mmꢁ1
1584
Crystal size
0.20 ꢂ 0.25 ꢂ 0.30 mm
h Range for data collection
Limiting indices
Reflections collected/unique
Completeness to h = 29.96
Refinement method
Data/restraints/parameters
Goodness-of-fit on F2
Final R indices [I > 2r(I)]
R indices (all data)
Absolute structure parameter
Largest diff. peak and hole
2.12–29.96°
ꢁ18 6 h 6 18, ꢁ23 6 k 6 23, ꢁ25 6 l 6 18
60393/21734 Rint = 0.0600
93.5%
Full-matrix least squares on F2
21734/2/958
bridge Crystallographic Database Centre as
Publication No. CCDC 679753.
a Supplementary
0.990
R1 = 0.0528, wR2 = 0.1095
R1 = 0.2036, wR2 = 0.1357
0.3(7)
2.3. NMR measurements
0.676 and ꢁ0.356 e Åꢁ3
The NMR spectra were recorded in CDCl3 using a Varian Gemini
300 MHz spectrometer. All spectra were locked to deuterium reso-
nance of CDCl3. The error in parts per million values was 0.01. All
1H NMR measurements were carried out at the operating fre-
quency 300.075 MHz; flip angle, pw = 45o; spectral width,
sw = 4500 Hz; acquisition time, at 2.0 s; relaxation delay,
d1 = 1.0 s; T = 293.0 K and TMS as the internal standard. No win-
dow function or zero filling was used. Digital resolution was
0.2 Hz/point. 13C NMR spectra were recorded at the operating fre-
quency 75.454 MHz; pw = 60o; sw = 19,000 Hz; at 1.8 s; d1 = 1.0 s;
T = 293.0 K and TMS as the internal standard.
The 1H and 13C NMR signals were assigned independently for
each species using one- or two-dimensional (COSY and HETCOR)
spectra.
2.4. FT-IR measurements
In the mid-infrared region the FT-IR spectra of NAFA were re-
corded in chloroform solution and potassium bromide as pellets.
A cell with Si windows and wedge-shaped layers was used to avoid

´
483
A. Huczynski et al. / Journal of Molecular Structure 891 (2008) 481–490
Table 2 (continued)
Table 2
Atomic coordinates (ꢂ104) and equivalent isotropic displacement parameters
x
y
z
U(eq)
56(1)
(Å2 ꢂ 103), (U(eq) is defined as one-third of the trace of the orthogonalized Uij tensor)
C(140)
C(150)
C(160)
C(170)
C(180)
C(190)
C(200)
C(210)
C(220)
C(230)
C(240)
C(250)
C(260)
C(270)
C(280)
C(290)
C(300)
C(310)
C(320)
C(330)
C(340)
C(350)
C(360)
C(370)
C(380)
C(390)
C(400)
C(410)
C(420)
C(430)
C(440)
C(450)
1225(2)
1780(2)
1318(2)
2237(2)
3115(3)
4089(2)
4107(2)
5167(2)
5983(2)
5871(2)
6180(3)
7029(2)
7932(3)
3279(6)
3504(5)
455(3)
9565(2)
9579(2)
9137(2)
9014(2)
8865(2)
9224(2)
3125(2)
3925(2)
4501(2)
5084(2)
4645(2)
4994(2)
5068(2)
5399(2)
5000(2)
4924(2)
5625(2)
5385(2)
5054(2)
4617(3)
4003(4)
4779(2)
2887(2)
2135(2)
2135(2)
3351(2)
1382(2)
2904(2)
2425(2)
1924(2)
1879(2)
1387(2)
927(2)
x
y
z
U(eq)
79(1)
57(1)
70(1)
72(1)
60(1)
59(1)
69(1)
74(1)
63(1)
62(1)
98(1)
83(1)
96(1)
209(4)
229(4)
107(2)
71(1)
78(1)
77(1)
89(1)
72(1)
80(1)
64(1)
65(1)
80(1)
96(1)
107(2)
101(1)
107(2)
101(2)
84(1)
75(1)
O(1)
O(2)
O(3)
O(4)
O(5)
O(6)
O(7)
O(8)
C(1)
C(2)
C(3)
C(4)
C(5)
C(6)
C(7)
C(8)
ꢁ495(2)
4847(1)
4971(2)
5859(2)
5227(1)
4418(2)
4195(1)
3983(1)
5424(1)
5215(2)
5936(2)
6227(3)
6884(3)
7276(3)
7031(3)
6369(2)
6220(2)
6502(2)
6506(2)
5701(2)
5697(2)
4856(2)
4604(2)
4581(2)
4162(2)
3995(2)
3826(2)
4194(2)
5081(2)
5337(2)
5044(2)
4156(2)
3640(2)
5277(2)
5000(3)
2946(3)
2535(5)
4654(3)
3851(2)
3983(3)
6193(2)
6919(2)
7165(3)
4152(2)
3467(2)
2948(2)
3033(2)
2525(3)
1886(3)
1760(3)
2197(3)
2695(4)
3343(3)
2297(3)
9816(1)
9917(1)
2988(1)
ꢁ2147(2)
ꢁ3123(2)
1330(2)
3373(2)
2372(2)
230(2)
ꢁ676(2)
ꢁ1365(4)
ꢁ1316(3)
ꢁ2230(4)
ꢁ2275(5)
ꢁ1393(5)
ꢁ484(4)
ꢁ423(3)
573(3)
2608(2)
3359(2)
1772(1)
2552(1)
879(1)
112(1)
116(1)
70(1)
86(1)
68(1)
60(1)
76(1)
73(1)
63(1)
85(1)
97(2)
110(2)
98(1)
74(1)
71(1)
61(1)
63(1)
59(1)
58(1)
64(1)
62(1)
59(1)
71(1)
82(1)
66(1)
66(1)
72(1)
86(1)
70(1)
65(1)
86(1)
93(1)
120(2)
116(2)
295(5)
102(1)
89(1)
142(2)
82(1)
79(1)
156(2)
80(1)
72(1)
67(1)
77(1)
99(1)
109(2)
106(2)
121(2)
123(2)
102(1)
80(1)
70(1)
89(1)
77(1)
62(1)
75(1)
57(1)
60(1)
70(1)
61(1)
53(1)
57(1)
60(1)
73(1)
75(1)
59(1)
66(1)
64(1)
60(1)
51(1)
49(1)
56(1)
10101(2)
10384(2)
10067(2)
9187(2)
8720(2)
10312(2)
10002(2)
7938(4)
7799(3)
9575(3)
8810(2)
8865(2)
11126(2)
11854(2)
12118(2)
9085(2)
8390(2)
8114(2)
8466(2)
8190(3)
7578(3)
7217(3)
7091(3)
7362(3)
8006(3)
7467(2)
581(1)
995(1)
2949(2)
3404(2)
3608(2)
4036(3)
4266(2)
4072(2)
3629(2)
3367(2)
2559(2)
2283(2)
2263(2)
2032(2)
2062(2)
1435(2)
728(2)
668(2)
23(2)
C(9)
535(2)
1819(2)
3984(3)
9372(2)
6426(3)
6524(3)
5688(3)
4712(3)
3929(3)
4062(4)
4961(4)
6748(4)
7542(4)
7412(3)
5790(3)
C(10)
C(11)
C(12)
C(13)
C(14)
C(15)
C(16)
C(17)
C(18)
C(19)
C(20)
C(21)
C(22)
C(23)
C(24)
C(25)
C(26)
C(27)
C(28)
C(29)
C(30)
C(31)
C(32)
C(33)
C(34)
C(35)
C(36)
C(37)
C(38)
C(39)
C(40)
C(41)
C(42)
C(43)
C(44)
C(45)
O(10)
O(20)
O(30)
O(40)
O(50)
O(60)
O(70)
O(80)
C(10)
C(20)
C(30)
C(40)
C(50)
C(60)
C(70)
C(80)
C(90)
C(100)
C(110)
C(120)
C(130)
1547(2)
2032(2)
3077(2)
3481(2)
4045(2)
3266(2)
3552(2)
2529(3)
1851(3)
788(2)
60(2)
ꢁ367(2)
214(2)
8(2)
938(2)
1521(3)
2008(3)
2457(2)
1453(2)
733(3)
ꢁ111(2)
ꢁ271(2)
310(2)
ꢁ368(3)
ꢁ986(3)
ꢁ832(2)
ꢁ1362(3)
ꢁ2097(3)
ꢁ2782(3)
1845(4)
1217(6)
4252(3)
4655(3)
5553(3)
3829(2)
1434(3)
ꢁ3273(4)
ꢁ447(3)
ꢁ101(3)
ꢁ779(3)
ꢁ1820(3)
ꢁ2463(4)
ꢁ2101(4)
ꢁ1101(5)
642(5)
409(2)
ꢁ233(2)
83(2)
630(2)
435(3)
interferences (mean layer thickness 170 lm). The spectra were ta-
ken with an IFS 113v FT-IR spectrophotometer (Bruker, Karlsruhe)
equipped with a DTGS detector; resolution 2 cmꢁ1, NSS = 64. The
Happ-Genzel apodization function was used.
ꢁ142(5)
ꢁ347(2)
1637(2)
2242(2)
2524(2)
1540(2)
4235(3)
2529(2)
2993(2)
3262(2)
3114(2)
3364(2)
3771(3)
3941(2)
3817(3)
3584(3)
3159(3)
3690(2)
2477(1)
2862(1)
2240(1)
3173(1)
2180(1)
3929(1)
4577(1)
4285(1)
2481(2)
2002(2)
1905(2)
1460(2)
1131(2)
1233(2)
1661(2)
1786(2)
2519(2)
2633(2)
2617(2)
2706(2)
2635(2)
2.5. UV–vis and fluorescence measurements
The absorption spectra of LAS and NAFA were recorded in 1 cm
cells on a spectrophotometer Jasco V-550 (JASCO, Tokyo, Japan).
The UV–vis spectra of LAS and NAFA (1 ꢂ 10ꢁ5 mol dmꢁ3) were re-
corded in spectral grade. The intensity of fluorescence emission
was measured for LAS and NAFA at the angle 90° relative to the
incident excitation light. During the absorption and fluorescence
measurements the samples were kept at room temperature. The
samples were excited at 260 nm. The fluorescence quantum yield
of NAFA was estimated from the equation:
1300(4)
909(4)
ꢀ
ꢁ
2
AR FS nS
ꢁ386(4)
6431(2)
8083(2)
9133(2)
4152(2)
2227(2)
2772(1)
4872(2)
5919(2)
7307(3)
7292(2)
8246(2)
8337(3)
7468(3)
6528(3)
6409(2)
5346(2)
5151(2)
4058(2)
3575(2)
2457(2)
2007(2)
US ¼ UR ꢀ
ꢀ
FR
AS
nR
where UR is the quantum yield of the standard, A is the absorbance
at the wavelength of the excitation, F is the integrated intensity of
the emission spectra, n is the refractive index of solution, the index
(S) means sample and (R) means reference. The standard fluoro-
phore for the quantum yield measurements was Quinine sulphate
in 0.2 mol dmꢁ3 H2SO4 (UR = 0.51) [26].
10852(1)
10122(1)
9336(1)
9236(1)
8997(1)
10412(1)
10171(2)
10862(2)
11178(2)
11817(2)
12144(2)
11882(2)
11247(2)
11044(2)
11384(2)
11405(2)
10601(2)
10604(2)
9784(2)
2.6. Semiempirical calculations
PM5 semiempirical calculations were performed using the
WinMopac 2003 program. In all cases full-geometry optimisation
of NAFA was carried out without any symmetry constraints [27–30].
2.7. Elemental analysis
The elemental analysis of NAFA was carried out on Vario ELIII
(Elementar, Germany).

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A. Huczynski et al. / Journal of Molecular Structure 891 (2008) 481–490
484
2.8. Melting point
11778, Enterococcus hirae ATCC 10541, Micrococcus luteus ATCC
9341, M. luteus ATCC 10240; Gram-negative rods: Escherichia coli
ATCC 10538, E. coli ATCC 25922, E. coli NCTC 8196, Proteus vulgaris
NCTC 4635, Pseudomonas aeruginosa ATCC 15442, P. aeruginosa
NCTC 6749, P. aeruginosa ATCC 27853, Bordetella bronchiseptica
ATCC 4617 and yeasts: Candida albicans ATCC 10231, C. albicans
ATCC 90028, Candida parapsilosis ATCC 22019.
Other micro-organisms which we used were obtained from the
Department of Pharmaceutical Microbiology, Medical University of
Warsaw, Poland. Antimicrobial activity was examined by the disc-
Corrected melting point was determined on a Mettler FP900.
2.9. Microbiological analysis
Micro-organisms used in this study were as follows: Gram-po-
sitive cocci: Staphylococcus aureus NCTC 4163, S. aureus ATCC
25923, S. aureus ATCC 6538, S. aureus ATCC 29213, S. epidermidis
ATCC 12228, Bacillus subtilis ATCC 6633, Bacillus cereus ATCC
Fig. 1. (a) A perspective view of NAFA in the crystal structure. Hydrogen bonds have been indicated by dashed lines; (b) The structures of NAFA without the engagement of
the C@O carbonyl group in hydrogen bond (type A), calculated by semiempirical PM5 method (WinMopac 2003).

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A. Huczynski et al. / Journal of Molecular Structure 891 (2008) 481–490
Table 3
diffusion method under standard conditions using Mueller–Hinton
Bond lengths (Å) and angles (°)
II agar medium (Becton–Dickinson) for bacteria and RPMI agar
with 2% glucose (Sigma) according to CLSI (previously NCCLS)
guidelines [31]. Sterile filter paper discs (9 mm diameter, What-
man No. 3 chromatography paper) were contaminated with the
compound tested solutions (in MeOH or MeOH/DMSO 1:1) to load
400 lg of a given compound per disc. Dry discs were placed on the
surface of appropriate agar medium. The results (diameter of the
growth inhibition zone) were read after 18 h of incubation at
35 °C. The compounds which showed activity in the disc-diffusion
tests were further examined by the agar dilution method to deter-
mine their MIC – minimal inhibitory concentration (CLSI) [32].
Concentrations of the agents tested in solid medium ranged from
3.125 to 400 lg mlꢁ1. The final inoculum of all organisms studied
was 104 CFUmlꢁ1 (colony forming units per millilitre), except the
I
II (‘)
O(1)AC(1)
O(1)AC(35)
O(2)AC(1)
1.319(8)
1.469(7)
1.212(8)
1.370(9)
1.452(6)
1.205(7)
1.436(6)
1.463(7)
1.439(6)
1.444(7)
1.431(7)
1.490(9)
1.415(9)
1.429(9)
1.383(10)
1.370(11)
1.521(11)
1.388(11)
1.409(10)
1.515(9)
1.567(7)
1.524(8)
1.526(8)
1.534(8)
1.529(8)
1.513(8)
1.536(8)
1.544(8)
1.541(9)
1.545(8)
1.532(8)
1.502(8)
1.543(8)
1.543(8)
1.552(8)
1.559(11)
1.533(8)
1.526(8)
1.542(9)
1.537(8)
1.539(9)
1.561(8)
1.547(9)
1.44(2)
1.323(7)
1.481(7)
1.242(7)
1.374(7)
1.445(6)
1.210(6)
1.454(6)
1.479(7)
1.450(6)
1.432(7)
1.441(7)
1.476(8)
1.417(8)
1.427(8)
1.387(8)
1.351(8)
1.509(8)
1.380(8)
1.370(8)
1.520(8)
1.539(7)
1.515(7)
1.518(8)
1.554(8)
1.535(7)
1.540(7)
1.525(8)
1.537(8)
1.522(8)
1.553(7)
1.518(8)
1.523(8)
1.535(8)
1.554(8)
1.498(8)
1.603(12)
1.507(8)
1.537(8)
1.515(8)
1.533(8)
1.517(8)
1.528(8)
1.535(8)
1.245(15)
1.528(7)
1.503(9)
1.355(9)
1.422(9)
1.433(9)
1.429(9)
1.365(9)
1.380(11)
1.353(11)
1.415(10)
1.367(11)
1.426(10)
1.408(11)
O(3)AC(3)
O(4)AC(11)
O(5)AC(13)
O(6)AC(15)
O(6)AC(18)
O(7)AC(19)
O(7)AC(23)
O(8)AC(22)
C(1)AC(2)
C(2)AC(7)
C(2)AC(3)
C(3)AC(4)
C(4)AC(5)
C(4)AC(34)
C(5)AC(6)
C(6)AC(7)
C(7)AC(8)
C(8)AC(9)
C(9)AC(10)
C(10)AC(11)
C(10)AC(33)
C(11)AC(12)
C(12)AC(32)
C(12)AC(13)
C(13)AC(14)
C(14)AC(30)
C(14)AC(15)
C(15)AC(16)
C(16)AC(17)
C(16)AC(29)
C(17)AC(18)
C(18)AC(19)
C(18)AC(27)
C(19)AC(20)
C(20)AC(21)
C(21)AC(22)
C(22)AC(25)
C(22)AC(23)
C(23)AC(24)
C(25)AC(26)
C(27)AC(28)
C(30)AC(31)
C(35)AC(36)
C(36)AC(44)
C(36)AC(37)
C(37)AC(38)
C(37)AC(45)
C(38)AC(39)
C(39)AC(40)
C(40)AC(41)
C(41)AC(45)
C(42)AC(43)
C(42)AC(45)
C(43)AC(44)
final inoculum for E. hirae ATCC 10541, which was 105 CFU mlꢁ1
.
Minimal inhibitory concentrations were read after 18 h of incuba-
tion at 35 °C.
3. Results and discussion
The structures and the numbering of the atoms of NAFA are
shown in Scheme 1.
3.1. X-ray structure analysis
NAFA crystallizes as a non-solvate in acetonitrile solution. The
molecular dimensions (Table 3) fully confirm the chemical formula
of the substance. Three intramolecular hydrogen bonds are charac-
teristic of all the NAFA structures. The O(3)AH(3)ꢀ ꢀ ꢀO(2) bond is
very short (Table 4) most likely due to the overcrowding effect of
the two adjacent substituents in a rigid fragment of conjugated
p-electron bonds. The other hydrogen bonds, between
1.532(9)
1.485(9)
1.360(10)
1.416(9)
1.389(9)
1.421(9)
1.356(10)
1.375(11)
1.348(11)
1.456(11)
1.347(12)
1.374(11)
1.412(12)
C(1)AO(1)AC(35)
C(15)AO(6)AC(18)
C(19)AO(7)AC(23)
O(2)AC(1)AO(1)
O(2)AC(1)AC(2)
O(1)AC(1)AC(2)
C(7)AC(2)AC(3)
C(7)AC(2)AC(1)
C(3)AC(2)AC(1)
O(3)AC(3)AC(4)
O(3)AC(3)AC(2)
C(4)AC(3)AC(2)
C(5)AC(4)AC(3)
C(5)AC(4)AC(34)
C(3)AC(4)AC(34)
C(4)AC(5)AC(6)
C(5)AC(6)AC(7)
C(6)AC(7)AC(2)
118.2(6)
109.8(4)
113.2(4)
123.5(8)
122.6(8)
113.8(8)
117.5(7)
124.2(7)
118.3(8)
116.9(9)
119.6(8)
123.4(9)
117.6(9)
121.4(10)
121.0(10)
121.6(9)
121.7(9)
118.0(7)
117.5(5)
109.6(4)
113.9(4)
120.6(6)
123.6(7)
115.8(6)
118.2(6)
125.0(6)
116.7(6)
116.0(6)
121.4(6)
122.5(6)
116.4(6)
124.0(6)
119.6(6)
123.2(7)
122.0(6)
117.5(6)
Fig. 2. The structures of NAFA with the engagement of the C@O carbonyl group in
hydrogen bond (type B) calculated by the semiempirical PM5 method (WinMopac
2003).
(continued on next page)

´
A. Huczynski et al. / Journal of Molecular Structure 891 (2008) 481–490
486
Table 3 (continued)
ened by hydrogen bonds, exhibits considerable differences in con-
formation. In particular, the position of the naphthalene
substituent is rotated by ca. 180° in molecule I relative to that in
molecule II (Fig. 1). Also conformational differences are found for
the ether bridge in the near fragment (Table 5), and for the ethyl
substituent at C(18). There are no strong intermolecular hydrogen
bonds in this crystal structure (Fig. 3). Therefore, it is plausible that
the molecular conformation of the fragment in which intramolec-
ular H-bonds are located is primarily stabilized by intramolecular
interactions, whereas the overall molecular conformation is signif-
icantly influenced by intermolecular interactions of the molecules
in the crystal. It has been recently shown that the range of inter-
molecular interactions in molecular crystals can be considerably
longer than the sums of van der Waals radii [33–35], as postulated
by Dance [36]. Evidence was also provided that intermolecular
interactions at various thermodynamical conditions can consider-
ably change the hydrogen bonding pattern in crystal structures
[37–38], or they can be responsible for stabilisation of specific
compounds [39–40].
I
II (‘)
C(6)AC(7)AC(8)
C(2)AC(7)AC(8)
C(7)AC(8)AC(9)
C(10)AC(9)AC(8)
117.2(8)
124.5(7)
110.8(5)
115.1(5)
114.2(5)
109.5(5)
112.1(5)
110.9(5)
106.3(5)
115.1(5)
112.5(5)
107.7(5)
109.1(5)
121.8(6)
121.7(6)
116.5(6)
110.8(6)
116.7(6)
107.4(5)
105.3(5)
108.8(5)
118.5(5)
101.0(5)
115.0(6)
112.8(6)
105.0(5)
104.5(5)
109.5(5)
111.8(6)
102.5(6)
113.1(7)
114.5(6)
109.6(5)
106.2(5)
117.4(6)
109.6(6)
112.4(6)
105.2(6)
109.1(5)
113.3(6)
111.0(6)
109.2(6)
109.1(6)
109.8(5)
111.7(6)
115.5(6)
113.1(6)
107.7(10)
112.5(7)
109.9(5)
119.7(8)
117.8(8)
122.6(7)
123.2(8)
117.8(8)
119.0(8)
122.7(8)
120.5(9)
120.6(9)
120.5(9)
123.9(10)
118.0(10)
121.4(9)
118.0(9)
124.1(10)
117.8(8)
117.8(6)
124.5(6)
109.9(5)
115.7(5)
113.4(5)
108.8(5)
112.4(5)
110.4(4)
108.7(5)
114.7(5)
112.1(5)
107.6(5)
111.7(5)
120.4(6)
121.5(6)
118.1(6)
112.6(5)
115.5(5)
106.8(5)
105.6(5)
107.6(5)
118.4(5)
101.2(5)
113.5(5)
113.9(6)
104.3(5)
104.1(5)
110.0(5)
112.9(5)
114.8(6)
107.3(6)
107.7(7)
108.3(5)
108.4(5)
116.6(5)
110.5(5)
112.8(5)
103.6(5)
109.3(5)
112.7(6)
111.0(6)
110.8(5)
109.4(6)
109.9(5)
112.1(6)
115.5(6)
116.3(6)
105.8(11)
113.3(6)
110.1(5)
117.9(8)
121.0(8)
121.1(7)
121.3(8)
117.6(7)
121.0(7)
119.7(8)
121.1(9)
122.5(9)
118.6(8)
120.3(9)
120.1(9)
122.8(8)
117.8(8)
121.8(9)
120.4(8)
C(9)AC(10)AC(11)
C(9)AC(10)AC(33)
C(11)AC(10)AC(33)
O(4)AC(11)AC(12)
O(4)AC(11)AC(10)
C(12)AC(11)AC(10)
C(32)AC(12)AC(11)
C(32)AC(12)AC(13)
C(11)AC(12)AC(13)
O(5)AC(13)AC(12)
O(5)AC(13)AC(14)
C(12)AC(13)AC(14)
C(30)AC(14)AC(13)
C(30)AC(14)AC(15)
C(13)AC(14)AC(15)
O(6)AC(15)AC(16)
O(6)AC(15)AC(14)
C(16)AC(15)AC(14)
C(17)AC(16)AC(15)
C(17)AC(16)AC(29)
C(15)AC(16)AC(29)
C(16)AC(17)AC(18)
O(6)AC(18)AC(17)
O(6)AC(18)AC(19)
C(17)AC(18)AC(19)
O(6)AC(18)AC(27)
C(17)AC(18)AC(27)
C(19)AC(18)AC(27)
O(7)AC(19)AC(20)
O(7)AC(19)AC(18)
C(20)AC(19)AC(18)
C(21)AC(20)AC(19)
C(20)AC(21)AC(22)
O(8)AC(22)AC(25)
O(8)AC(22)AC(23)
C(25)AC(22)AC(23)
O(8)AC(22)AC(21)
C(25)AC(22)AC(21)
C(23)AC(22)AC(21)
O(7)AC(23)AC(22)
O(7)AC(23)AC(24)
C(22)AC(23)AC(24)
C(22)AC(25)AC(26)
C(28)AC(27)AC(18)
C(31)AC(30)AC(14)
O(1)AC(35)AC(36)
C(44)AC(36)AC(37)
C(44)AC(36)AC(35)
C(37)AC(36)AC(35)
C(38)AC(37)AC(36)
C(38)AC(37)AC(45)
C(36)AC(37)AC(45)
C(39)AC(38)AC(37)
C(38)AC(39)AC(40)
C(41)AC(40)AC(39)
C(40)AC(41)AC(45)
C(43)AC(42)AC(45)
C(42)AC(43)AC(44)
C(36)AC(44)AC(43)
C(42)AC(45)AC(37)
C(42)AC(45)AC(41)
C(37)AC(45)AC(41)
3.2. NMR measurements
The 1H and 13C NMR data of NAFA are collected in Table 6. In
the 1H NMR spectrum of NAFA in CDCl3 solution the most signif-
icantly shifted signal is found at 11.52 ppm. This signal is as-
signed to the donor proton of the O(3)H group at the benzene
ring which is involved the O(3)Hꢀ ꢀ ꢀO(2)@C intramolecular hydro-
gen bond. The signal of the O(4)H and O(8)H protons arises as
one broadened signal at 3.56 ppm. The position of this signal
indicates that both protons are involved in relatively weak
hydrogen bonds.
The signals of naphthalene ring protons are found in the range
between 7.44 and 8.07 ppm, whereas the C(5)H and C(6)H proton
signals of the salicylic ring arise as two doublets at 7.13 and
6.61 ppm, respectively (Fig. 4). The signal of the methylene group
proton is observed at 5.84 ppm as a double doublets due to the
geminal (2J) spin–spin coupling (Fig. 4). This kind of coupling indi-
cates that both protons are located in different electronic environ-
ments and a typical AB spin–spin coupling structure is observed.
The most interesting signals in the 13C NMR spectrum of NAFA
are those of C(1) and C(3) carbon atoms which suggest the exis-
tence of enol and/or ketol tautomeric forms of the lasalocid esters.
These signals arise at 171.90 and 160.88 ppm, respectively, indicat-
ing that NAFA exists in solution predominantly in the enol form as
shown in Scheme 1.
Table 4
The parameters (length and angle) of intramolecular hydrogen bonds of NAFA
determined from X-ray studies and calculated by PM5 semiempirical method
Used method Compound
Atoms engaged
Length (Å) Angle (°)
in hydrogen bonds
X-ray
PM5
NAFA(type
O(3)Hꢀ ꢀ ꢀO(2)
O(8)Hꢀ ꢀ ꢀO(4)
O(4)Hꢀ ꢀ ꢀO(6)
O(30)Hꢀ ꢀ ꢀO(20)
O(80)Hꢀ ꢀ ꢀO(40)
O(40)Hꢀ ꢀ ꢀO(60)
O(3)Hꢀ ꢀ ꢀO(2)
O(8)Hꢀ ꢀ ꢀO(4)
O(4)Hꢀ ꢀ ꢀO(6)
O(3)Hꢀ ꢀ ꢀO(2)
O(8)Hꢀ ꢀ ꢀO(4)
O(4)Hꢀ ꢀ ꢀO(5)
2.556(8)
2.870(6)
2.922(6)
2.525(6)
2.869(6)
2.916(6)
149
164
177
148
165
169
A
- crystal)
NAFA(type
NAFA(type
2.59
2.85
2.93
2.60
2.89
2.85
146
161
173
145
150
128
A)
B)
O(8)Hꢀ ꢀ ꢀO(4)H and O(4)Hꢀ ꢀ ꢀO(6) are considerably longer. The
hydrogen bonds O(4)Hꢀ ꢀ ꢀO(6) and O(8)Hꢀ ꢀ ꢀO(4) are probably
responsible for the conformation of the molecular fragment con-
tained between C(11)OH and the terminal C(22)OH. Indeed, differ-
ences between the torsion angles of two symmetry-independent
molecules I and II in these fragments are smaller than few degrees
(Table 5). The opposite molecular moiety, which is not strength-
(Type A) – structure of NAFA in which the C(13)@O group is not engaged in
hydrogen bond, (type B) – structure of NAFA in which the C(13)@O group is
engaged in hydrogen bond.

´
487
A. Huczynski et al. / Journal of Molecular Structure 891 (2008) 481–490
Table 5 (continued)
Table 5
Selected torsion angles (°)
I
II
I
II
C(25)AC(22)AC(23)AC(24)
49.4(8)
50.7(8)
C(35)AO(1)AC(1)AO(2)
C(35)AO(1)AC(1)AC(2)
O(2)AC(1)AC(2)AC(7)
O(1)AC(1)AC(2)AC(7)
O(2)AC(1)AC(2)AC(3)
O(1)AC(1)AC(2)AC(3)
C(7)AC(2)AC(3)AO(3)
C(1)AC(2)AC(3)AO(3)
C(1)AC(2)AC(3)AC(4)
7.1(10)
ꢁ3.7(9)
C(21)AC(22)AC(23)AC(24)
O(8)AC(22)AC(25)AC(26)
C(23)AC(22)AC(25)AC(26)
C(21)AC(22)AC(25)AC(26)
O(6)AC(18)AC(27)AC(28)
C(17)AC(18)AC(27)AC(28)
C(19)AC(18)AC(27)AC(28)
C(13)AC(14)AC(30)AC(31)
C(15)AC(14)AC(30)AC(31)
C(1)AO(1)AC(35)AC(36)
O(1)AC(35)AC(36)AC(44)
O(1)AC(35)AC(36)AC(37)
C(35)AC(36)AC(37)AC(38)
C(35)AC(36)AC(37)AC(45)
C(35)AC(36)AC(44)AC(43)
ꢁ72.4(7)
ꢁ61.4(8)
57.6(9)
ꢁ73.0(7)
ꢁ63.1(8)
54.8(8)
ꢁ176.6(5)
ꢁ160.3(7)
23.4(9)
17.7(10)
ꢁ158.6(6)
175.5(6)
ꢁ2.6(10)
179.0(7)
ꢁ177.6(7)
1.5(11)
180.0(7)
ꢁ178.5(7)
171.8(7)
ꢁ178.6(6)
ꢁ170.1(6)
7.9(11)
175.3(5)
ꢁ168.4(6)
12.6(9)
179.4(7)
171.6(9)
ꢁ76.5(11)
53.1(11)
ꢁ69.2(8)
167.5(6)
ꢁ122.2(7)
ꢁ87.9(7)
92.3(7)
177.7(6)
28.5(13)
143.7(10)
ꢁ94.4(11)
ꢁ64.6(7)
172.3(5)
ꢁ93.3(7)
102.8(7)
ꢁ77.2(7)
ꢁ2.6(9)
8.8(9)
ꢁ170.2(5)
177.7(5)
0.3(8)
178.9(6)
ꢁ179.7(5)
ꢁ1.1(8)
O(3)AC(3)AC(4)AC(5)
O(3)AC(3)AC(4)AC(34)
C(2)AC(3)AC(4)AC(34)
C(34)AC(4)AC(5)AC(6)
C(5)AC(6)AC(7)AC(8)
C(1)AC(2)AC(7)AC(6)
C(3)AC(2)AC(7)AC(8)
ꢁ179.8(5)
ꢁ177.1(6)
175.4(6)
179.9(6)
ꢁ172.2(6)
5.0(10)
0.1(10)
178.7(6)
ꢁ178.5(7)
178.1(6)
ꢁ178.7(6)
C(1)AC(2)AC(7)AC(8)
C(6)AC(7)AC(8)AC(9)
C(2)AC(7)AC(8)AC(9)
ꢁ100.5(7)
ꢁ97.2(7)
73.1(8)
77.7(8)
C(7)AC(8)AC(9)AC(10)
C(8)AC(9)AC(10)AC(11)
C(8)AC(9)AC(10)AC(33)
C(9)AC(10)AC(11)AO(4)
C(33)AC(10)AC(11)AO(4)
C(9)AC(10)AC(11)AC(12)
C(33)AC(10)AC(11)AC(12)
O(4)AC(11)AC(12)AC(32)
C(10)AC(11)AC(12)AC(32)
O(4)AC(11)AC(12)AC(13)
C(10)AC(11)AC(12)AC(13)
C(32)AC(12)AC(13)AO(5)
C(11)AC(12)AC(13)AO(5)
C(32)AC(12)AC(13)AC(14)
C(11)AC(12)AC(13)AC(14)
O(5)AC(13)AC(14)AC(30)
C(12)AC(13)AC(14)AC(30)
O(5)AC(13)AC(14)AC(15)
C(12)AC(13)AC(14)AC(15)
C(18)AO(6)AC(15)AC(16)
C(18)AO(6)AC(15)AC(14)
C(30)AC(14)AC(15)AO(6)
C(13)AC(14)AC(15)AO(6)
C(30)AC(14)AC(15)AC(16)
C(13)AC(14)AC(15)AC(16)
O(6)AC(15)AC(16)AC(17)
C(14)AC(15)AC(16)AC(17)
O(6)AC(15)AC(16)AC(29)
C(14)AC(15)AC(16)AC(29)
C(15)AC(16)AC(17)AC(18)
C(29)AC(16)AC(17)AC(18)
C(15)AO(6)AC(18)AC(17)
C(15)AO(6)AC(18)AC(19)
C(15)AO(6)AC(18)AC(27)
C(16)AC(17)AC(18)AO(6)
C(16)AC(17)AC(18)AC(19)
C(16)AC(17)AC(18)AC(27)
C(23)AO(7)AC(19)AC(20)
C(23)AO(7)AC(19)AC(18)
O(6)AC(18)AC(19)AO(7)
C(17)AC(18)AC(19)AO(7)
C(27)AC(18)AC(19)AO(7)
O(6)AC(18)AC(19)AC(20)
C(17)AC(18)AC(19)AC(20)
C(27)AC(18)AC(19)AC(20)
O(7)AC(19)AC(20)AC(21)
C(18)AC(19)AC(20)AC(21)
C(19)AC(20)AC(21)AC(22)
C(20)AC(21)AC(22)AO(8)
C(20)AC(21)AC(22)AC(25)
C(20)AC(21)AC(22)AC(23)
C(19)AO(7)AC(23)AC(22)
C(19)AO(7)AC(23)AC(24)
O(8)AC(22)AC(23)AO(7)
C(25)AC(22)AC(23)AO(7)
C(21)AC(22)AC(23)AO(7)
O(8)AC(22)AC(23)AC(24)
170.1(6)
62.7(7)
165.5(5)
61.0(7)
3.3. FT-IR measurements
ꢁ170.6(5)
ꢁ173.2(6)
In Fig. 5a and b, the spectra of NAFA in chloroform (dashed line)
and in KBr (solid line) are compared. In the spectrum of NAFA in
KBr only one band assigned to the m(OH) vibrations of O(4)H and
O(8)H groups is observed at 3447 cmꢁ1 indicating that these OH
groups form relatively weak hydrogen bonds of comparable
strength. This observation is in good agreement with the X-ray
structure of NAFA. The vibration of the proton in the intramolecu-
lar OAHꢀꢀꢀO@C hydrogen bond, which is the strongest of all hydro-
gen bonds should be observed in the FT-IR spectrum in the region
below 3000 cmꢁ1. However, this absorption does neither appear
very clearly in the NAFA solid state spectrum nor in the spectrum
of chloroform solution. The reason for the low intensity of this
absorption is probably a strong coupling of the proton motion in
the intramolecular hydrogen bond with the p electrons, which
eventually results in a loss of proton polarizability [41].
In the spectrum of NAFA in chloroform solution a broad band
with a maximum about 3451 cmꢁ1 and a new weak one at about
3580 cmꢁ1 are observed (Fig. 5b, dashed line). The latter band indi-
cates that in chloroform solution one of the weak hydrogen bonds
is partially broken. This indicates that the structure of NAFA under-
go slight changes in solution as compared to the solid state. A sim-
ilar conclusion can be drawn from the spectra in the region of the
m(C@O) vibrations (Fig. 5b). It is obvious, that the NAFA spectrum of
the solid state (solid line) exhibits only one band at 1714 cmꢁ1 as-
signed to the m(C@O) vibrations of C(13)@O group. However, in a
chloroform solution the intensity of this band decreases and an
additional shoulder at 1702 cmꢁ1 arises. The existence of the
shoulder at 1702 cmꢁ1 means that in a chloroform solution the ke-
tone C(13)@O group becomes partially hydrogen bonded, which
supports the above statement that the solution structure of NAFA
is slightly different relative to that in the solid state.
60.3(6)
57.8(6)
ꢁ65.0(6)
ꢁ176.5(5)
58.2(7)
179.3(5)
58.6(7)
ꢁ61.2(6)
178.1(5)
81.5(7)
ꢁ66.2(6)
ꢁ178.2(5)
57.9(7)
178.6(5)
55.5(7)
ꢁ60.5(6)
176.3(5)
83.3(6)
ꢁ40.1(8)
ꢁ93.2(6)
143.4(5)
ꢁ9.2(8)
ꢁ41.0(8)
ꢁ97.9(6)
139.7(5)
ꢁ15.1(9)
164.2(5)
113.4(7)
ꢁ67.3(7)
ꢁ24.4(6)
ꢁ152.3(5)
79.9(7)
ꢁ45.1(7)
ꢁ40.2(8)
ꢁ165.3(6)
36.7(6)
158.6(6)
160.0(5)
ꢁ78.1(8)
ꢁ35.1(7)
ꢁ156.8(6)
1.9(7)
ꢁ118.0(5)
120.1(6)
21.7(7)
140.0(6)
ꢁ89.0(7)
63.1(6)
ꢁ169.2(5)
ꢁ65.7(6)
178.9(5)
48.7(8)
167.2(5)
118.6(6)
ꢁ64.9(6)
ꢁ23.6(6)
ꢁ151.0(5)
85.7(6)
ꢁ40.4(6)
ꢁ33.8(8)
ꢁ159.9(6)
37.3(6)
157.9(6)
159.5(5)
ꢁ79.9(8)
ꢁ36.8(7)
ꢁ159.3(6)
0.0(6)
ꢁ121.3(5)
117.0(7)
23.5(6)
142.7(6)
ꢁ98.7(7)
63.0(6)
ꢁ169.7(5)
ꢁ68.9(6)
175.3(5)
56.9(7)
Furthermore, the band characteristic of the ester group at about
1650 cmꢁ1 seems to be unchanged in both states.
57.3(7)
53.5(7)
ꢁ58.1(8)
171.7(6)
ꢁ56.0(7)
ꢁ177.2(5)
52.4(8)
ꢁ62.3(8)
179.4(6)
ꢁ54.7(6)
ꢁ177.2(5)
51.8(8)
3.4. UV–vis and fluorescence studies
The absorption and emission spectra of NAFA are shown in
Fig. 6 and the spectral characteristics are given in Table 7. In the
absorption spectrum of NAFA an intense band at 226 nm together
with less intense satellite bands in the region 250–350 nm are ob-
served (Fig. 6 left, solid line). The absorption maximum at 281 nm
in UV–vis spectrum of NAFA is caused by p–p* transitions in the
naphthalene ring. The respective spectrum of lasalocid acid is less
structured. Only three bands with maxima at 215, 248 and 318 nm
(Fig. 6, dashed line) are observed. The fluorescence spectrum of
68.3(8)
69.9(8)
ꢁ176.2(6)
ꢁ51.9(8)
ꢁ62.8(6)
66.6(7)
ꢁ66.4(6)
176.8(5)
55.0(7)
ꢁ175.6(6)
ꢁ50.8(7)
ꢁ63.5(6)
66.3(7)
ꢁ66.8(6)
178.6(5)
54.9(6)
166.2(5)
165.3(5)

´
A. Huczynski et al. / Journal of Molecular Structure 891 (2008) 481–490
488
NAFA shows a band with a maximum at ca. 373 nm (kexc = 260 nm)
(Fig. 6 right, solid line). In the fluorescence emission spectrum of
lasalocid acid this band is shifted to 381 nm (kexc = 260 nm)
(Fig. 6 right, dashed line). The quantum yield of the fluorescence
emission of lasalocid acid was calculated to be U = 0.15, which is
quite high. Note that the salicylic acid moiety of the lasalocid mol-
ecule actively contributes to the emission [42]. In contrast, the new
1-naphthylmethyl ester of lasalocid acid shows a three orders of
magnitude lower quantum yield of fluorescence (Table 7), which
indicates an influence of the carboxylic group on this process.
semiempirical method on the basis of the crystallographic data
(Fig. 1b). This comparison clearly shows that both structures are
very similar despite the fact that these two methods describe
two different states, i.e. the gas and the solid states. This result
means that the PM5 semiempirical method is a reliable tool to
determine and visualise three-dimensional structures of chemical
compounds also in the solid state.
In Fig. 2, we show type B structure, which is obtained by PM5
semiempirical calculations on the basis of our spectroscopic data
in solution. In contrast to type A structure, the C(13)@O(5) keto
group forms an intramolecular hydrogen bond, which is neither
observed in the crystals nor in the PM5 semiempirical calcula-
tions based on the crystallographic data (see Fig. 1a and b). This
hydrogen bond and two further intramolecular hydrogen bonds
stabilize type B structure of NAFA and permit distinction of
two parts of the ester molecule. One part including the salicylic
group is stabilized by the O(3)AHꢀ ꢀ ꢀO(2)@C intramolecular
hydrogen bond and the other part of the molecule is stabilized
by the O(8)AHꢀ ꢀ ꢀO(4) and O(4)AHꢀ ꢀ ꢀO(5)@C intramolecular
hydrogen bonds.
3.5. PM5 calculations
On the basis of our spectroscopic and X-ray results, two alterna-
tive structures of NAFA, labelled types A and B are calculated by the
PM5 semiempirical method. In type A structure the C(13)@O(5)
keto group is not involved in a hydrogen bond. Interestingly, this
type of structure is found in the crystals as revealed by X-ray anal-
ysis. In Fig. 1, we compare the crystal structure obtained by X-ray
analysis (Fig. 1a) to the calculated structure, obtained by the PM5
Fig. 3. Arrangement of complex NAFA in the crystal lattice, projected along the [100] crystal direction.

´
489
A. Huczynski et al. / Journal of Molecular Structure 891 (2008) 481–490
Table 6
1H NMR and 13C NMR chemical shifts (ppm) of NAFA in CDCl3
100
3451
No. of atom
1H NMR
13C NMR
1
2
3
4
5
6
7
8
9
–
–
–
–
171.90
110.05
160.88
123.98
135.20
121.72
144.67
34.04
35.98
34.04
71.40
49.25
215.18
54.77
85.02
35.08
39.22
85.88
73.13
20.91
29.33
70.73
76.86
14.02
29.88
6.42
3580
50
0
7.13
6.61
–
2.71
1.32 1.48
1.46
3.43
2.68
–
2.72
3.75
2.21
1.45 1.82
–
3.66
1.45 1.68
1.38 1.54
–
3.76
1.18
1.58
0.88
1.27
0.82
1.02
1.38, 1.82
0.88
0.43
0.58
2.21
5.84
–
3447
3500
4000
3000
2500
2000
1500
1000
500
-1
Wavenumber [cm
]
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
O(3)H
O(4)H
O(8)H
100
50
0
1711
1650
1702
1714
1750
1800
1700
1650
-1
1600
1550
Wavenumber [cm
]
30.63
8.49
Fig. 5. FT-IR spectra of NAFA: (—) in KBr pallet, (---) in chloroform: (a) 4000–
400 cmꢁ1; (b) 1800–1550 cmꢁ1
.
16.01
18.24
12.04
13.17
12.68
15.87
65.67
133.71
130.63
123.55
126.87
126.12
128.70
129.75
125.29
128.53
131.81
–
1.0
0.5
0.0
1.0
0.5
0.0
–
8.06
7.54
7.49
7.87
7.87
7.47
7.62
–
11.52
3.56
3.56
–
–
200
250
300
350
400
450
500
Wavelength [nm]
The calculated hydrogen bond parameters of both NAFA
structures together with the respective X-ray data are collected in
Table 4.
Fig. 6. Absorption (left) and emission corrected spectra (right) of: NAFA (—), and
LAS (—).
Fig. 4. 1H NMR spectrum of NAFA in region of aromatic proton signals.

´
A. Huczynski et al. / Journal of Molecular Structure 891 (2008) 481–490
490
Table 7
sis). Moreover, as expected, the cell walls of Gram-negative bacte-
ria do not permit the penetration of hydrophobic molecules with
high weights and thus the micro-organisms are not susceptible
to the action of lasalocid as well its ester.
UV–vis and fluorescence spectral characteristics of NAFA
UV–vis
Fluorescence
kexc (nm)
260
kmax (nm)
log e
kem (nm)
U
226.0
281.0
317.0
4.38
3.87
3.67
373
2.9 ꢂ 10ꢁ4
Acknowledgement
´
Adam Huczynski thanks the Foundation for Polish Science for
fellowship.
kmax,
maximum of the absorption band; e, molar extinction coefficient (dmꢁ3
molꢁ1 cmꢁ1), kexc, excitation wavelength; kem, maximum of the fluorescence band; U,
quantum yield of fluorescence relative to quinine sulphate in 0.1 N H2SO4 [26].
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inhibitory concentration (MIC) [lg/ml]
LAS
GIZ
NAFA
GIZ
MIC
MIC
S. aureus NCTC 4163
S. aureus ATCC 25923
S. aureus ATCC 6538
S. aureus ATCC 29213
S. epidermidis ATCC 12228
B. subtilis ATCC 6633
B. cereus ATCC 11778
E. hirae ATCC 10541
M. luteus ATCC 9341
M. luteus ATCC 10240
E. coli ATCC 10538
E. coli ATCC 25922
E. coli NCTC 8196
P. vulgaris NCTC 4635
P. aeruginosa ATCC 15442
P. aeruginosa NCTC 6749
P. aeruginosa ATCC 27853
B. bronchiseptica ATCC 4617
C. albicans ATCC 10231
C. albicans ATCC 90028
C. parapsilosis ATCC 22019
35
29
31
30
27
28
28
20
26
32
na
na
na
na
na
na
na
na
na
na
na
12.5
12.5
12.5
12.5
12.5
6.25
6.25
12.5
12.5
12.5
19
15
19
14
13
14
18
Marks
Marks
14
na
na
na
na
na
na
>400
>400
>400
>400
>400
400
100
>400
>400
400
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na
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3.6. Antimicrobial activity
Lasalocid acid (LAS) and its new 1-naphthylmethyl ester (NAFA)
were tested in vitro for their antibacterial and antifungal activity.
We used the following microorganisms in this study: Gram-posi-
tive bacteria, Gram-negative rods and yeasts (Table 8). The antimi-
crobial properties of both LAS and NAFA are characterised by the
minimum inhibitory concentration (MIC). The results are pre-
sented in Table 8 [31,32]. Lasalocid acid and its 1-naphthylmethyl
ester exhibit activity against Gram-positive bacteria. The activity of
Lasalocid towards against the human pathogenic bacteria (MIC
6.25–12.5 lg/ml) is higher as compared to its ester derivative
(MIC 100 to >400 lg/ml). Both compounds tested (LAS and NAFA)
are inactive against strains of Candida (C. albicans and C. parapsilo-
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