Structure reactivity and thermodynamic analysis on the oxidation of ampicillin drug by copper(III) complex in aqueous alkaline medium (stopped-flow technique)

Article abstract of DOI:10.1016/j.molstruc.2009.05.013

Oxidation of penicillin derivative, ampicillin (AMP) by diperiodatocuprate(III) (DPC) in alkaline medium at a constant ionic strength of 0.01-mol dm^-3 was studied spectrophotometrically. The reaction between DPC and ampicillin in alkaline mediu

Full text of DOI:10.1016/j.molstruc.2009.05.013

Journal of Molecular Structure 930 (2009) 180–186
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Journal of Molecular Structure
journal homepage: www.elsevier.com/locate/molstruc
Structure reactivity and thermodynamic analysis on the oxidation of ampicillin
drug by copper(III) complex in aqueous alkaline medium (stopped-flow technique)
*
Nagaraj P. Shetti, Rajesh N. Hegde, Sharanappa T. Nandibewoor
P.G. Department of Studies in Chemistry, Karnatak University, Pavate Nagar, Dharwad 580 003, India
a r t i c l e i n f o
a b s t r a c t
Article history:
Oxidation of penicillin derivative, ampicillin (AMP) by diperiodatocuprate(III) (DPC) in alkaline medium
at a constant ionic strength of 0.01-mol dmꢀ3 was studied spectrophotometrically. The reaction between
DPC and ampicillin in alkaline medium exhibits 1:4 stoichiometry (ampicillin:DPC). Intervention of free
radicals was observed in the reaction. Based on the observed orders and experimental evidences, a mech-
anism involving the protonated form of DPC as the reactive oxidant species has been proposed. The oxi-
dation reaction in alkaline medium has been shown to proceed via a DPC–AMP complex, which
decomposes slowly in a rate determining step to yield phenyl glycine (PG) and free radical species of
6-aminopenicillanic acid (6-APA), followed by other fast steps to give the products. The two major prod-
ucts were characterized by IR, NMR, LC–MS and Spot test. The reaction constants involved in the different
steps of the mechanism were calculated. The activation parameters with respect to slow step of the
mechanism were computed and discussed and thermodynamic quantities were also determined.
Ó 2009 Elsevier B.V. All rights reserved.
Received 10 February 2009
Received in revised form 24 April 2009
Accepted 6 May 2009
Available online 13 May 2009
Keywords:
Oxidation
Mechanism
Ampicillin
Diperiodatocuprate(III)
Thermodynamic studies
1. Introduction
place in oxidation chemistry due to their abundance and relevance
in biological chemistry [13,14]. When the copper(III) periodate
Semi-synthetic b-lactam antibiotics are the most important
class of antibacterial agents. Ampicillin (AMP), (6R)-6-( -phenyl-
-glycylamino)penicillanic acid is a semi-synthetic penicillin. Pen-
complex is oxidant and multiple equilibria between different cop-
per(III) species are involved, it would be interesting to know which
of the species is the active oxidant.
a
D
icillin-G was the first antibiotic to be used in the chemotherapy. It
is a bacteriostatic and a drug of choice for the treatment of the
infections caused by most of the Gram positive and Gram-negative
bacteria [1]. Ampicillin is a broad spectrum antibiotic. Ampicillin,
due to its acidic nature inhibits the protein synthesis of the bacte-
rial cell wall. The basic nucleus of the ampicillin is 6-aminopenic-
illanic acid, which consists of a thiazolidine ring linked to b-lactam
ring. The side chain determines the antibacterial and pharmacolog-
ical characteristics of this compound [2]. The complexes of ampi-
cillin with different metal ions [3–5], the degradation of
ampicillin [6] and its spectrophotometric determination [7] have
been studied.
Periodate and tellurate complexes of copper in its trivalent state
have been extensively used in the analysis of several organic com-
pounds [8]. The kinetics of self-decomposition of these complexes
was studied in some detail [9]. Copper(III) is shown to be an inter-
mediate in the copper(II) catalysed oxidation of amino acids by
peroxydisulphate [10]. The oxidation reaction usually involves
the copper(II)–copper(I) couple and such aspects are detailed in
different reviews [11,12]. Copper complexes have occupied a major
As in the earlier literature [15], ampicillin is not stable in aque-
ous media. To study the structure reactivity on the oxidation of
ampicillin in alkali media, such stability becomes important and
interesting. Since, there is no report in the literature on kinetics
of oxidation of ampicillin drug by any oxidant in alkaline medium,
such oxidation studies may throw some light on the mechanism of
conversions of the compounds in biological systems. Connection of
stopped-flow technique with UV–visible spectroscopy is an effec-
tive method to detect the spectra of intermediatory species for
understanding the reaction mechanism. In earlier reports [16] on
DPC oxidation, periodate had retarding effect and the order in
[OH]ꢀ was found to be less than unity in most of the reactions.
However, in the present study, entirely different kinetic observa-
tions were obtained. Hence the title reaction is undertaken to
understand a suitable mechanism and active species in such
media.
2. Experimental
2.1. Materials and reagents
All chemicals used were of reagent grade and millipore water
was used throughout the work. A solution of ampicillin (Sigma–Al-
* Corresponding author. Tel.: +91 836 2215286; fax: +91 836 274788.
E-mail address: stnandibewoor@yahoo.com (S.T. Nandibewoor).
0022-2860/$ - see front matter Ó 2009 Elsevier B.V. All rights reserved.
doi:10.1016/j.molstruc.2009.05.013

N.P. Shetti et al. / Journal of Molecular Structure 930 (2009) 180–186
181
drich) was prepared by dissolving an appropriate amount of
recrystallised sample in millipore water. The purity of AMP sample
was checked by comparing its melting point 207 °C with the liter-
ature (208 °C) and the LC–MS spectra of ampicillin shows the
molecular ion peak, m/z at 348.7. The copper(III) periodate com-
plex was prepared and standardized [17,18]. UV–visible spectrum
with maximum absorption at 415 nm verified existence of cop-
per(III) complex. The solution of copper sulphate (BDH) was pre-
pared by dissolving the known amount of samples in millipore
water. Periodate solution was prepared and used after keeping it
for 24 h to complete the equilibrium. Its concentration was ascer-
tained iodometrically [19] at neutral pH by phosphate buffer. Re-
quired alkalinity and ionic strength were maintained by KOH
(BDH) and KNO3 (AR), respectively, in reaction solutions. t-Butyl
alcohol (S.D. Fine) was used to study the dielectric constant effect.
tion. No significant difference in the results was obtained under a
N2 atmosphere and in the presence of air. In view of the ubiquitous
contamination of carbonate in the basic medium, the effect of car-
bonate was also studied. Added carbonate had no effect on the
reaction rates. The spectral changes during the reaction are shown
in Fig. S1. It is evident from the figure that the concentration of DPC
through the absorbance decreases at 415 nm.
2.3. Instruments used
(a) For kinetic measurements: Varian CARY 50 Bio UV–visible
Spectrophotometer connected to a rapid kinetic accessory (HI-
TECH SFA-12).
(b) For product analysis: LC–ESI–MS, Hewlett Packard 1100 re-
verse phase HPLC system with a phenomenas C18 column, hp 1100
series diode array UV/Visible detector and hp 1100 MSD series
mass analyzer (Germany), Nicolet 5700-FT-IR spectrometer (Ther-
mo USA), 300 MHz 1H NMR and 13C NMR spectrometer (Brucker,
Switzerland), The pH of the medium in the solution was measured
by ELICO (LI613) pH meter.
2.2. Kinetic measurements
Since the ampicillin and DPC reaction was fast, the kinetic mea-
surements were performed on a Varian CARY 50 Bio UV–visible
spectrophotometer attached to a rapid kinetic accessory (HI-TECH
SFA-12). The kinetics was followed under pseudo-first order condi-
tion where [AMP] > [DPC] at 32.0 0.1 °C, unless specified. The pH
of reaction medium was maintained in the range of 10–11. The
reaction was initiated by mixing the DPC to AMP solution which
also contained required concentration of KNO3, KOH and KIO4.
The progress of reaction was followed spectrophotometrically at
415 nm by monitoring the decrease in absorbance due to DPC with
3. Results
3.1. Stoichiometry and product analysis
Different sets of reaction mixtures containing varying ratios of
DPC to ampicillin in presence constant amounts of OHꢀ and
KNO3 were kept for 2 h in closed vessel under nitrogen atmo-
sphere. The remaining concentration of DPC was estimated by
spectrophotometrically at 415 nm. The results indicated 1:4 stoi-
chiometry as given in Scheme 1. The stoichiometry was also con-
firmed by Job’s method (Fig. 1) (Table S1).
the molar absorbancy index, ‘
(literature = 6230 [20]). It was verified that there is a negligible
e
’ to be 6235 100 dm3 molꢀ1 cmꢀ1
e
interference from other species present in the reaction mixture
at this wavelength.
The reaction was followed to more than 95% completion of reac-
tion. Plots of log (absorbance) versus time lead to the first order
rate constants (kobs). The plots were linear up to 85% completion
of the reaction and rate constants were reproducible with in 5%.
The main two products, oxophenyl acetic acid and 2-formyl-
5,5-dimethyl-thiazolidine-4-carboxlic acid were identified by TLC
and separated by column chromatography over neutral alumina
using a mixture of benzene and chloroform (80:20 v/v) as eluent.
Further evaporation of the each fractions gave a pure product,
which showed a single spot on TLC plate.
IR spectrum of oxophenyl acetic acid showed a sharp absorption
peak at 1744 cmꢀ1 (due to ketone C@O stretch), 1648 cmꢀ1 (due to
carboxylic C@O stretch) and a broad peak at 3341 cmꢀ1 (due to
carboxylic OH).
During
the
kinetics
a
constant
concentration
viz.
5.0 ꢁ 10ꢀ5 mol dmꢀ3 of KIO4 was used throughout the study unless
otherwise stated. Since periodate is present in the excess in DPC,
the possibility of oxidation of ampicillin by periodate in alkaline
medium at 32 °C was tested. The progress of the reaction was fol-
lowed iodometrically. However, it was found that there was no sig-
nificant reaction under the experimental conditions employed
compared to the DPC oxidation of ampicillin. The total concentra-
tions of periodate and OHꢀ was calculated by considering the
amount present in the DPC solution and that additionally added.
Kinetics runs were also carried out in N2 atmosphere in order to
understand the effect of dissolved oxygen on the rate of the reac-
Further this product was characterized by LC–MS, which gave
m/z at 149 (m ꢀ 1). % yield = 42%; elemental analysis found % (calcd
% for C8H8O3) C, 64.05 (64.11); H, 4.03 (3.95). An acidic solution of
2,4-dinitrophenylhydrazine was added, and after few minutes a
red precipitate appeared, indicating the presence of
[21,22].
a-keto acids
NH
2
O
H
-
COO
4 [OH-]
S
CH
3
NH
+ 4 [Cu(OH)2(H3IO6)2]3-
+ Cu(OH)2 +
CH
3
O
N
O
-
1
S
COO
CH
CH
3
3
2
5
O
C
H
HN
-
COO
4
3
+ 8 [H3IO6]2- +CO2 + 2NH3 + H+
.................... (1)
Scheme 1. Stoichiometry of oxidation of ampicillin by alkaline diperiodatocuprate(III).

182
N.P. Shetti et al. / Journal of Molecular Structure 930 (2009) 180–186
Table 1
0.03
0.02
0.01
0.00
Effect of [DPC], [AMP] and [OHꢀ] on diperiodatocuprate(III) oxidation of ampicillin in
alkaline medium at 32 °C, I = 0.01 mol dmꢀ3
.
[DPC] ꢁ 105
[AMP] ꢁ 104
[OHꢀ] ꢁ 102
kobs ꢁ 102ðsꢀ1
Þ
(mol dmꢀ3
)
(mol dmꢀ3
)
(mol dmꢀ3
)
Found Calculated
1.0
3.0
5.0
7.0
10.0
5.0
5.0
5.0
5.0
5.0
0.5
0.5
0.5
0.5
0.5
9.39
9.13
9.22
9.42
9.23
9.48
9.48
9.48
9.48
9.48
5.0
5.0
5.0
5.0
5
2.0
5.0
8.0
12.0
16.0
20.0
0.5
0.5
0.5
0.5
0.5
0.5
4.46
9.22
12.7
16.4
18.5
20.5
4.55
9.48
12.9
16.4
18.8
20.6
0.08
0.10
0.12
0.14
0.16
0.18
0.20
Mole fraction of DPC
5.0
Fig. 1. Job’s plot using absorbance values measured at the end of the reaction at
32 °C (conditions as mentioned in Table S1).
5.0
5.0
5.0
5.0
5.0
5.0
5.0
5.0
5.0
5.0
0.1
0.3
0.5
0.8
1.0
15.8
12.7
9.22
7.39
6.11
16.4
12.0
9.48
7.20
6.20
The IR spectrum of 2-formyl-5,5-dimethyl-thiazolidine-4-car-
boxylic acid showed a sharp absorption peak at 1742 cmꢀ1 (due
to aldehyde C@O stretch), 1668 cmꢀ1 (due to carboxylic C@O
stretch) and 1384 cmꢀ1 (due to ACH3 stretch). NAH stretching
indicated the order with respect to [DPC] was unity (Table 1). This
was also confirmed by linearity of the plots of log [absorbance] ver-
sus time (r P 0.998, S < 0.002) up to 85% completion of the reaction
(Fig. 2).
showed absorptions at 3437 cmꢀ1
.
Further this product was characterized by LC–MS, which gave
m/z at 190 (m + 1). % yield = 28%; elemental analysis found % (calcd
% for C7H11NO3S) C, 44.43 (44.51); H, 5.86 (5.81); N, 7.40 (7.35); S,
16.95 (16.89). Both the products were also identified by preparing
their 2,4 DNP derivatives.
3.5. Effect of [ampicillin]
The by-products were identified as ammonia by Nessler’s re-
agent [23] and the CO2 was qualitatively detected by bubbling
nitrogen gas through the acidified reaction mixture and passing
the liberated gas through tube containing lime water.
The effect of ampicillin on the rate of reaction was studied at
constant concentrations of alkali, DPC and periodate at a constant
ionic strength of 0.01 mol dmꢀ3. The substrate, ampicillin was var-
ied in the range of 2.0 ꢁ 10ꢀ4–2.0 ꢁ 10ꢀ3 mol dmꢀ3. The kobs values
increased with increase in concentration of ampicillin. The order
with respect to [ampicillin] was found to be less than unity (Table
1) (r P 0.999, S < 0.004). This is also confirmed in the plot of kobs
versus [AMP]0.66 which is linear rather than the direct plot of kobs
versus [AMP] (Fig. 3).
3.2. 1H NMR and 13C NMR studies
The products oxophenyl acetic acid and 2-formyl-5,5-dimethyl-
thiazolidine-4-carboxylic acid were characterized by 1H and also
13C NMR spectra through differently positioned proton and carbon.
Oxophenyl acetic acid: 1H NMR (300 MHz, DMSO-d6) d: 10.88
(s, 1H, carboxylic OH exchangeable with D2O), 7.42–7.78 (m, 5H,
ArH). 13C NMR (75 MHz, DMSO-d6) d: 180.3 and 164.4 (carboxylic
and keto carbon, respectively), 136.4, 134.2, 129.8, 129.7, 129.1,
129.2 for aromatic carbon.
3.6. Effect of [alkali]
The effect of increase in concentration of alkali on the rate of
reaction was studied at constant concentrations of ampicillin,
DPC and periodate at a constant ionic strength of 0.01 mol dmꢀ3
at 32 °C. The rate constants decreased with increase in alkali con-
2-Formyl-5,5-dimethyl-thiazolidine-4-carboxylic acid: 1H NMR
(300 MHz, CDCl3-d6) d: 1.62 (s, 6H, gem dimethyl), 2.63 (s, 1H, NH
exchangeable with D2O), 4.05 (s, 1H, 4CAH), 4.31 (s, 1H, 2CAH), 9.1
(s, 1H, formyl), 10.12 (s, 1H, carboxylic OH exchangeable with
D2O). 13C NMR (75 MHz, CDCl3-d6) d: 183.6 and 174.5 (aldehyde
and carboxylic carbons) 81.3, 74.8, 42.1, 26.8 and 26.7 for methyl
carbons.
Regression analysis of experimental data to obtain the regres-
sion coefficient r and standard deviation S of point from the regres-
sion line was performed using Microsoft Excel programme.
3.0
2.5
2.0
1.5
1.0
(5)
(4)
3.3. Reaction orders
The reaction orders were determined from the slope of log kobs
versus log (concentration) plots by varying the concentrations of
ampicillin, alkali and periodate in turn while keeping all other con-
centrations and conditions constant, except DPC.
(3)
0.5
0.0
(2)
(1)
0
10
20
30
3.4. Effect of [diperiodatocuprate(III)]
Time in Sec
The oxidant DPC concentration was varied in the range of
Fig. 2. First order plots on the oxidation of ampicillin by DPC in aqueous alkaline
medium. [DPC] ꢁ 105 (mol dmꢀ3): (1) 1.0; (2) 3.0; (3) 5 .0; (4) 7.0; and (5) 10.0.
1.0 ꢁ 10ꢀ5–1.0 ꢁ 10ꢀ4 mol dmꢀ3 and the fairly constant kobs values

N.P. Shetti et al. / Journal of Molecular Structure 930 (2009) 180–186
183
kept for 2 h in an inert atmosphere. On diluting the reaction mix-
ture with methanol, a white precipitate was formed, indicating
the intervention of free radicals in the reaction. The blank experi-
ments of either DPC or ampicillin alone with acrylonitrile did not
induce any polymerization under the same condition as those in-
duced for reaction mixture. Initially added acrylonitrile decreased
the rate of reaction indicating free radical intervention, which is
the case in earlier work [24].
[AMP]0.66 x 102 mol dm-3
1.5 1.0 0.5
2.0
0.0
0.0
0.5
1.0
1.5
2.5
2.0
1.5
1.0
0.5
0.0
3.11. Effect of temperature
The kinetics was studied at six different temperatures (12, 17,
22, 27, 32 and 37 °C) under varying concentrations of ampicillin
and alkali keeping other conditions constant. The rate constants in-
creased with increase in temperature. The rate constants (k) of the
slow step of the reaction mechanism were obtained from the
slopes and intercepts of 1/kobs versus 1/[AMP] plot at six different
temperatures and were used to calculate the activation parame-
ters. The energy of activation corresponding to these constants
was evaluated from the Arrhenius plot of log k versus 1/T
(r P 0.9925, S < 0.011) and other activation parameters obtained
are tabulated in Table 2.
2.0
2.5
0
1
2
3
[AMP] x 103 mol dm-3
Fig. 3. Plot of kobs versus [AMP]0.66 and kobs versus [AMP].
centration (Table 1), indicating negative fractional order depen-
dence on alkali concentration (r P 0.992, S < 0.003). While varying
the alkali concentrations, the pH range was in between 10 and 11.
4. Discussion
In the later period of 20th century the kinetics of oxidation of
various organic and inorganic substrates have been studied by
Cu(III) species which may be due to its strong versatile nature of
one-electron oxidant. The water soluble copper(III) periodate com-
plex is reported [25] to be [Cu(HIO6)2(OH)2]7ꢀ. However, in an
aqueous alkaline medium and at a high pH range employed in
the study, periodate is unlikely to exist as HIO6 (as present in
the complex) as is evident from its involvement in the multiple
equilibria [26] Eqs. (2)–(4) depending on the pH of the solution.
3.7. Effect of [periodate]
The effect of increasing concentration of periodate was studied
by varying the periodate concentration from 5.0 ꢁ 10ꢀ5 to
5.0 ꢁ 10ꢀ4 mol dmꢀ3 keeping all other reactant concentrations
and conditions constant. It was found that the added periodate
had negligible effect on the rate of reaction.
4ꢀ
3.8. Effect of ionic strength (I) and dielectric constant of the medium
(D)
Table 2
The ionic strength of the reaction medium was varied by using
different amounts of KNO3 from 0.005 to 0.05 mol dmꢀ3, while the
concentrations all other reactants were held constant. The rate
constants increased with an increase in the ionic strength of the
reaction medium. The plot of log kobs versus I1/2 was linear with po-
sitive slope.
Varying the t-butyl alcohol and water percentage varied dielec-
tric constant of the medium, ‘D’. The D values were calculated from
the equation D = DwVw + DBVB, where Dw and DB are dielectric con-
stants of pure water and t-butyl alcohol, respectively, and Vw and
VB are the volume fractions of components water and t-butyl alco-
hol, respectively, in the total mixture. The decrease in dielectric
constant of the reaction medium increased the rate of the reaction.
Under our experimental conditions t-butyl alcohol–water mixture
solvent did not undergo oxidation by DPC. The plot of log kobs ver-
sus 1/D was linear with positive slope.
Thermodynamic activation parameters for the oxidation of ampicillin by DPC in
aqueous alkaline medium with respect to the slow step of Scheme 2.
Temperature (K)
k ꢁ 10ꢀ1 (sꢀ1
)
(A) Effect of temperature
285
290
295
300
305
310
1.12
1.67
1.94
2.55
3.39
3.91
Parameters
Values
(B) Activation parameters (Scheme 2)
D
D
D
H# (kJ molꢀ1
)
)
)
34.0 1.5
S# (JK molꢀ1
G# (kJ molꢀ1
ꢀ143 10
77
2
log A
6.0 0.2
Temperature (K)
K4 ꢁ 103 (mol dmꢀ3
)
K5 ꢁ 10ꢀ2 (dm3 molꢀ1
)
3.9. Effect of initially added products
(C) Effect of temperature to calculate K4 and K5 for the oxidation of ampicillin by
diperiodatocuprate(III) in alkaline medium
285
290
295
300
305
310
0.74 0.03
0.89 0.03
1.1 0.04
1.5 0.04
1.8 0.05
2.1 0.12
2.03 0.02
4.85 0.04
6.30 0.05
20.6 1.0
29.1 1.2
40.7 2.0
One of the products, copper(II) (CuSO4) in the concentration
range of 1.0 ꢁ 10ꢀ5–1.0 ꢁ 10ꢀ4 mol dmꢀ3 with all other reactant
concentrations and conditions kept constant, did not have any sig-
nificant effect on the rate of the reaction.
3.10. Polymerization study
Thermodynamic quantities
Values from K4
Values from K5
(D) Thermodynamic quantities using K4 and K5
H (kJ molꢀ1
S (JKꢀ1 molꢀ1
G305 (kJ molꢀ1
)
33
55
16 0.5
1
3
91
363 10
ꢀ20
5
The intervention of free radicals in the reaction was examined
by polymerization of acrylonitrile. The reaction mixture, to which
a known quantity of acrylonitrile monomer initially added, was
D
D
D
)
)
1

184
N.P. Shetti et al. / Journal of Molecular Structure 930 (2009) 180–186
K1
been obtained. The result of decrease in rate of reaction with in-
H5IO6 ¢ H4IOꢀ þ Hþ; K1 ¼ 5:1 ꢁ 10ꢀ4
ð2Þ
ð3Þ
ð4Þ
6
crease in alkalinity (Table 1) can be explained in terms of prevail-
ing equilibrium of formation of [Cu(OH)2(H3IO6)2]3ꢀ from
[Cu(OH)2(H3IO6)(H2IO6)]4ꢀ hydrolysis as given in the following
equation.
K2
H4IOꢀ ¢ H3IO2ꢀ þ Hþ; K2 ¼ 4:9 ꢁ 10ꢀ9
6
6
K3
H3IO2ꢀ ¢ H2IO3ꢀ þ Hþ; K3 ¼ 2:5 ꢁ 10ꢀ12
K4
½CuðOHÞ ðH3IO6ÞðH2IO6Þꢂ4ꢀ þ H2O ¢ ½CuðOHÞ ðH3IO6Þ ꢂ3ꢀ þ OHꢀ
6
6
2
2
2
ꢀ
Periodic acid exists as H5IO6 in acid medium and H4IO6 near
pH 7. Hence, under alkaline (pH 10.7) conditions as employed in
ð5Þ
2ꢀ
Such type of equilibrium Eq. (5) has been well noticed in the lit-
erature [27]. The less than unit order in [AMP] presumably results
from formation of a complex (C) between the protonated DPC spe-
cies and ampicillin prior to the formation of the products. This
complex (C) decomposes in a slow step and undergoes amide
hydrolysis, to form phenyl glycine and a free radical species de-
rived from ampicillin as intermediates. There are evidences for
the amide hydrolysis of ampicillin to form phenylglycine (PG)
and 6-aminopenicillanic acid (6-APA) in the literature [1]. Here
the presence of excess of alkali (pH 10.7) enhances nucleophilic at-
tack at b-lactum carbonyl carbon and causes opening of the unsta-
ble four membered b-lactum ring. The intermediate phenylglycine
then reacts with two more molecules of protonated DPC in a fur-
ther fast step; it undergoes oxidative deamination to form the
product such as oxophenyl acetic acid. The oxidative deamination
of amino acids are well documented in the literature [21,22]. The
free radical species of ampicillin further reacts with another mole-
this study, the main species are expected to be H3IO6
and
H2IO63ꢀ. Thus, at the pH employed in this study, the soluble cop-
per(III) periodate complex might be [Cu(OH)2(H3IO6)2]3ꢀ, a conclu-
sion also supported by earlier work [16].
The reaction between diperiodatocuprate(III) complex and
ampicillin in alkaline medium has the stoichiometry 1:4
(AMP:DPC) with a first order dependence on [DPC] and an apparent
order of less than unit order in [substrate], negligible effect of
added periodate and a negative fractional order dependence on [al-
kali]. No effect of added products was observed. Based on the
experimental results, a mechanism is proposed for which all the
observed orders in each constituent such as [oxidant], [reductant],
[OH]ꢀ and [IO4]ꢀ may be well accommodated.
In most of the reports [16] on DPC oxidation, OHꢀ had an
increasing effect on the rate of the reaction and periodate retarded
the reaction and MPC was considered as active species of DPC.
However, in the present kinetic study, different kinetic results have
K4
[Cu(OH)2(H3IO6)2]3- +OH-
[Cu(OH)2(H3IO6)(H2IO6)]4- + H2O
NH2
H
S
NH
K5
CH3
CH3
COO-
Complex (C)4-
+ [Cu(OH)2(H3IO6)2]3-
O
N
O
NH2
.
S
COO-
k / slow
2 OH-
H3N
-OOC
CH3
Complex (C)4-
+
+ Cu(OH)2 +
2[H3IO6]2-
HN
CH3
COO-
Intermediate (II)
Intermediate (I)
NH2
O
COO-
COO-
fast
2 [Cu(OH)2 (H3IO6)2]3-
+
+
OH-
+ NH4
+ 2 Cu(OH)2 + 4 [H3IO6]2-
Intermediate (I)
S
.
S
CH3
H3N
-OOC
CH3
HC
fast / OH-
-NH3, -CO2
+ [Cu(OH)2(H3IO6)2]3-
HN
CH3
CH3
O
HN
COO-
COO-
Intermediate (II)
+ Cu(OH)2 + 2 [H3IO6]2-
Scheme 2. Detailed scheme for the oxidation of ampicillin by alkaline diperiodatocuprate(III).

N.P. Shetti et al. / Journal of Molecular Structure 930 (2009) 180–186
185
cule of protonated DPC in a fast step; it undergoes decarboxylation
and deamination to form carboxylic aldehyde molecule, Cu(II) and
periodate species as given in Scheme 2.
1/[AMP] x 103dm3 mol-1
8
6
4
2
0
0
1
2
3
Since Scheme 2 is in accordance with the generally well-ac-
cepted principle of non-complementary oxidations taking place
in sequence of one-electron steps, the reaction between the sub-
strate and oxidant would afford a radical intermediate. This type
of radical scavenging experiment revealed such a possibility (see
infra). The direct plot of kobs versus [AMP] was drawn to know
the parallel reaction if any along with interaction of oxidant and
reductant. However, the plot of kobs versus [AMP] was not linear.
Thus, in Scheme 2, the parallel reaction and involvement of two
molecules of AMP in the complex are excluded. The probable struc-
ture of the complex (C) is given in Scheme 3 and it shows that the
carbonyl group of oxygen was coordinated to the metal complex as
in the literature [28].
1.5
1.0
0.5
0.0
Spectroscopic evidence for the complex formation between oxi-
dant and substrate was obtained from UV–visible spectra of ampi-
cillin (5 ꢁ 10ꢀ4), DPC (5.0 ꢁ 10ꢀ5), [OHꢀ] (0.005 mol dmꢀ3) and
mixture of both. A bathochromic shift of about 5 nm from 269 to
274 nm in the spectra of DPC was observed. The Michaelis–Menten
plot also proved the complex formation between DPC and ampicil-
lin, which explains the less than unit order dependence on [AMP].
Scheme 2 leads to the rate law Eq. (6)
0
0.4
0.8
1.2
[OH-] x 10-2 mol dm-3
Fig. 4. Verification of rate law (8) for the oxidation of ampicillin by diperiodato-
cuprate(III) at 32 °C.
1/T (r P 0.9604, S 6 0.006) and log K5 versus 1/T (r P 0.996,
S 6 0.007) and the values of enthalpy of reaction
D
H, entropy
ꢀd½DPCꢂ
kK4K5½AMPꢂ½DPCꢂ
of reaction
DS and free energy of reaction DG, were calculated
rate ¼
¼
ð6Þ
dt
½OHꢀꢂ þ K4 þ K4K5½AMPꢂ
for the first and second equilibrium steps. These values are given
in Table 2. A comparison of the thermodynamic quantities of
first step of Scheme 2 with those obtained for the slow step of
the reaction shows that these values mainly refer to the rate
limiting step, supporting the fact that the reaction before rate
determining step is fairly fast and involves low activation energy
[30].
rate
kK4K5½AMPꢂ
kobs
¼
¼
ð7Þ
½DPCꢂ ½OHꢀꢂ þ K4 þ K4K5½AMPꢂ
This explains all the observed kinetic orders of different species.
The rate law Eq. (7) can be rearranged in to the following form,
which is suitable for verification.
The effect of ionic strength and dielectric constant of medium
on the rate explains qualitatively the reaction between two nega-
tively charged ions, as seen in Scheme 2. The moderate values of
1
½OHꢀꢂ
1
1
k
¼
þ
þ
ð8Þ
kobs kK4K5½AMPꢂ kK5½AMPꢂ
D
H# and
D
S# were both favorable for electron transfer processes.
According to Eq. (8), other conditions being constant, plots of 1/
kobs versus [OHꢀ] (r P 0.998, S 6 0.014) and 1/kobs versus 1/[AMP]
(r P 0.997, S 6 0.016) should be linear and are found to be so
(Fig. 4). The slopes and intercepts of such plots lead to the values
The value of
DS
# within the range for radical reaction has been as-
cribed [31] to the nature of electron pairing and unpairing pro-
cesses and to the loss of degrees of freedom formerly available to
the reactants upon the formation of rigid transition state. The ob-
served modest enthalpy of activation and a relatively low value
of the entropy of activation as well as a higher rate constant of
the slow step indicate that the oxidation presumably occurs via in-
ner-sphere mechanism. This conclusion is supported by earlier
observation [32].
of
K4,
K5
and
k
as
(1.82 0.05) ꢁ 10ꢀ3 mol dmꢀ3
,
(29.1 1.2) ꢁ 102 dm3 molꢀ1 and (3.39 0.1) ꢁ 10 sꢀ1
, respec-
tively. The value of K4 is in good agreement with the earlier litera-
ture [29].
The thermodynamic quantities for the first, second and third
equilibrium steps of Scheme 2 can be evaluated as follows.
[AMP] and [OHꢀ] (as in Table 1) were varied at six different
temperatures. The plots of 1/kobs versus [OHꢀ] and 1/kobs versus
1/[AMP] should be linear (Fig. 4). From the slopes and intercepts,
the values of K4 and K5 were calculated at different temperatures
and these values are given in Table 2. The vant Hoff’s plots were
made for variation of K4 and K5 with temperature (log K4 versus
5. Conclusion
The simplified kinetic model is consistent and gives a reason-
able response to experimental data. Among various species of
DPC in alkaline medium, protonated DPC, [Cu(OH)2(H3IO6)2]3ꢀ is
considered as active species for the title reaction. The results indi-
cate that, the role of pH in the reaction medium is crucial. On oxi-
dation of ampicillin at higher pH, it undergoes amide hydrolysis
and gives the respective oxidized products. Rate constant of slow
step and other equilibrium constants of different steps, involved
in the mechanism were evaluated and activation parameters with
respect to slow step of reaction were computed. The overall mech-
anistic sequence described here is consistent with product studies,
mechanistic and kinetic studies.
4-
NH2
H
H
S
N
CH3
CH3
COO
OH
OH
OH
N
O
O
O
O
O
O
O
HO
HO
Cu
I
I
O
HO
OH
OH
Acknowledgments
One of the authors N.P.S. thanks to Dr. Namdev Shelke, Univer-
sity of Colorado, USA for his cooperation during this work and also
Scheme 3. Probable structure of the complex (C).

186
N.P. Shetti et al. / Journal of Molecular Structure 930 (2009) 180–186
thanks to Dr. S.K. Nataraj, Research Scientist, Chonnam National
University, Gwangju, Republic of Korea for his valuable
suggestions.
Appendix B. Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.molstruc.2009.05.013.
Appendix A
References
According to Scheme 2
[1] A.L.O. Ferreira, L.R.B. Goncalves, R.C. Giordano, R.L.C. Giordano, Braz. J. Chem.
Eng. 17 (2000) 4–7.
[2] J.P. Hou, J.W. Poole, J. Pharm. Sci. 58 (1969) 1510.
ꢀd½DPCꢂ
kK4K5½AMPꢂ½DPCꢂ
½OHꢀꢂ
rate ¼
¼ kðcÞ ¼
ðA-1Þ
[3] G. Mukherjee, T. Ghosh, J. Inorg. Biochem. 59 (1995) 827.
[4] V.G. Alekseev, I.S. Samuilova, Russ. J. Inorg. Chem. 53 (2008) 327.
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[9] G.I. Rozovoskii, A.K. Misyavichyus, A.Y. Prokopchik, Kinet. Catal. 16 (1975) 337.
[10] M.G. RamReddy, B. Sethuram, T. Navaneeth Rao, Ind. J. Chem. 16A (1978) 313.
[11] K.D. Karlin, Y. Gultneh, Progress in Inorganic Chemistry, in: S.J. Lipard (Ed.), vol.
35, Wiley, New York, 1997, p. 220.
[12] W.B. Tolman, Acc. Chem. Res. 30 (1997) 227.
[13] K.N. Kitajima, Y. Moro-oka, Chem. Rev. 94 (1994) 737.
[14] E.I. Solomon, P. Chen, M. Metz, S.K. Lee, A.E. Palmer, Angew. Chem. Int. Ed. Engl.
40 (2001) 4570.
[15] Y. Zhu, E. Roets, Z. Ni, M.L. Moreno, E. Porqueras, J. Hoogmartens, J. Pharm.
Bioanal. 14 (1996) 631, and references therein.
dt
½DPCꢂ ¼ ½DPCꢂ þ ½CuðOHÞ ðH3IO6Þ ꢂ3ꢀ þ ½Cꢂ
T
F
2
2
where T and F refer to total and free concentrations.
K4
K4K5½AMPꢂ
½OHꢀꢂ
½DPCꢂT ¼ ½DPCꢂF½1 þ
þ
½OHꢀꢂ
ðA-2Þ
ðA-3Þ
½DPCꢂT½OHꢀꢂ
½DPCꢂF ¼
½OHꢀꢂ þ K4 þ K4K5½AMPꢂ
Similarly,
[16] D.C. Hiremath, T.S. Kiran, S.T. Nandibewoor, Int. J. Chem. Kinet. 39 (2007) 236.
[17] P.K. Jaiswal, K.L. Yadava, Ind. J. Chem. 11 (1973) 83.
½AMPꢂT ¼ ½AMPꢂF þ ½Cꢂ
[18] C.P. Murthy, B. Sethuram, T. Navaneeth Rao, Z. Phys. Chem. 262 (1981) 336.
[19] G.P. Panigrahi, P.K. Misro, Ind. J. Chem. 16A (1978) 201.
[20] N.P. Shetti, S.T. Nandibewoor, Z. Phys. Chem. 223 (2009) 299.
[21] D. Laloo, M.K. Mohanti, J. Chem. Soc. Daltons Trans. (1990) 311.
[22] A.G. Raso, P.M. Deya, J.M. Saa, J. Org. Chem. 51 (1986) 4285.
[23] G.H. Jeffery, J. Bassett, R.C. Denney, Vogel’s Text Book of Quantitative Chemical
Analysis, fifth ed., ELBS, New York, 1996, p. 679.
[24] S. Bhattacharya, P. Banerjee, Bull. Chem. Soc. Jpn. 69 (1996) 3475.
[25] K.B. Reddy, B. Sethuram, T. Navneeth Rao, Z. Phys. Chem. 268 (1987) 706.
[26] J.C. Bailar Jr., H.J. Emeleus, S.R. Nyholm, A.F. Trotman Dikenson, Comprehensive
Inorganic Chemistry, vol. 2, Pergamon Press, Oxford, 1975, p. 1456.
[27] T.S. Kiran, D.C. Hiremath, S.T. Nandibewoor, Z. Phys. Chem. 221 (2007) 501.
[28] C. Orvig, M.J. Abrams, Chem. Rev. 99 (1999) 9.
½AMPꢂT½OHꢀꢂF
½AMPꢂF ¼
½OHꢀꢂ þ K4K5½DPCꢂ
F
F
In view of low concentrations of DPC used, compared to [OHꢀ], the
second term in the denominator is neglected.
½AMPꢂT ¼ ½AMPꢂF; similarly ½OHꢀꢂT ¼ ½OHꢀꢂF
ðA-4Þ
Substituting Eqs. (A-2) and (A-4), in Eq. (A-1) and omitting the sub-
scripts T and F we get
[29] S.D. Kulkarni, S.T. Nandibewoor, Transition Met. Chem. 31 (2006) 1034.
[30] K.S. Rangappa, M.P. Raghavendra, D.S. Mahadevappa, D. Channegouda, J. Org.
Chem. 63 (1998) 531.
[31] C. Walling, Free Radicals in Solution, Academic Press, New York 1957, p.38.
[32] S.A. Farokhi, S.T. Nandibewoor, Tetrahedron 59 (2003) 7595.
ꢀd½DPCꢂ
kK4K5½AMPꢂ½DPCꢂ
rate ¼
¼
ðA-5Þ
dt
½OHꢀꢂ þ K4 þ K4K5½AMPꢂ