Iranian Journal of Pharmaceutical Research (2013), 12 (4): 659-669 Copyright © 2013 by School of Pharmacy

Received: August 2012 Shaheed Beheshti University of Medical Sciences and Health Services
Accepted: April 2013

Original Article

Investigation of Behavior of Forced Degradation of Lidocaine HCl by

NMR Spectroscopy and GC-FID Methods: Validation of GC-FID Method
for Determination of Related Substance in Pharmaceutical Formulations
Yucel Kadioglua*, Alptug Atilaa, Mehmet Serdar Gultekinb, Nurdan Alcan Alpb

a
Department of Analytical Chemistry, Faculty of Pharmacy, Ataturk University, 25240,
Erzurum, Turkey. b Department of Chemistry, Faculty of Science, Ataturk University, 25240,
Erzurum, Turkey.

Abstract

The forced degradation study of lidocaine HCl was carried out according to the ICH
guideline Q1A (R2). The degradation conditions were assessed to be hydrolysis, oxidation,
photolysis and dry heat during 24 h, 48 h and 72 h and then the samples were investigated by GC-
FID method and nuclear magnetic resonance (NMR) spectroscopy. According to these results,
the degradation products were not observed in all reaction conditions during the 72 h period.
Only spectral changes in the 1H and 13C-NMR spectrum were observed in hydrogen peroxide
and acid degradation. As a result of this degradation, n-oxide was formed. After acid-induced
degradation with HCl, the secondary amine salt was formed. Furthermore, trifluoroacetic
acid (TFA) was used as the acidic media, and the decomposition products were observed. A
simple and reliable gas chromatography method with flame ionization detection (GC-FID) was
developed and validated for the determination of lidocaine HCl in pharmaceutical formulations
in the form of a cream and injections. The GC-FID method can be used for a routine analysis
of lidocaine HCl in pharmaceutical formulations and the proposed method, together with NMR
spectroscopy, can be applied in stability studies.

Keywords: GC-FID; NMR; Lidocaine HCl; Forced degradation.

Introduction fenil)-etanamid hydrochloride (Figure 1 A), is

a local anesthetic substance of the amide type
Local anesthesia is described as any technique (2). The analysis of lidocaine is important for
that induces the absence of sensation in part of keeping abreast of the patient’s medical process
the body, generally for the aim of inducing local and quality controlling studies in pharmaceutical
analgesia, that is, local insensitivity to pain, formulations. Because the anesthetic action of
although other local senses may be affected as the local anesthetic substances is well known and
well (1, 2). appears to be especially useful on skin damaged
In 1943, lidocaine was first synthesized under by sun exposure, local anesthetic substances like
the name Xylocaine by Swedish chemist Nils lidocaine can be illegally present in cosmetic
Löfgren as an alternative class of anesthetics. products, particularly in the formulation of after-
Lidocaine HCl, 2-diethylamino-N-(2,6-dimethyl sun cosmetics (2, 3).
The literature shows that the thin-layer
* Corresponding author: chromatography method (4), electroanalytical
E-mail: [email protected] method (5), atomic absorption spectrophotometry
Kadioglu Y et al. / IJPR (2013), 12 (4): 659-669

Figure 1. Chemical structure of lidocaine HCl (A) and IS [prilocaine HCl (B)].

method (6, 7), spectrophotometric methods (8- vasoconstrictors in local anesthetic solutions
13), HPLC methods with different detection (10, (lidocaine and mepivacaine) (34), the short-
13-19), capillary electrophoretic method (20) and term stability of lidocaine-adrenaline epidural
membrane electrode method (21, 22) have been solution in pH-adjusted solutions (35), the
reported for determination of lidocaine HCl in chemical stability of bupivacaine, lidocaine and
pharmaceutical formulations. The determination epinephrine in pH-adjusted solutions (36), the
of mixtures with other local anesthetics of stability of lidocaine and oxycodone in rectal
lidocaine HCl in biological samples (23-28) and gel with the HPLC method (37), the stability of
pharmaceutical formulation (29) have been done lidocaine and epinephrine solutions exposed to
with the GC-MS and GC methods. There are electric current (38), the stability of lidocaine
only a few studies for determination of lidocaine HCl and lidocaine HCl with epinephrine
HCl with the GC method in whole blood (30), injectable solutions with the HPLC method
serum (31), and urine (32). (39), the stability with gradient HPLC-DAD of
According to the current good manufacturing miconazole nitrate and lidocaine HCl in their
practices, the stability-indicating assay method combined oral gel dosage form (40) and finally in
before release must be assessed for all drugs. A accordance with the study of George et al., who
forced degradation study of the drug substance exposed the four local anesthetics (lidocaine,
can help to identify the possible degradation mepivacaine, procaine and prilocaine) to
products, which can help to establish the standard autoclaving conditions (250 °C and 20
degradation pathways and the intrinsic stability - 30 min) and analyzed the solutions with the gas
of the molecule and validate the stability- chromatography method (41).
indicating power of the analytical procedures The aim of the present work is to develop and
used (33). The nature of the forced degradation validate a new GC-FID method for determination
will depend on the individual drug substance and of lidocaine HCl in pharmaceutical formulations
the type of the drug product involved (33). with a simple sample preparation using internal
So far, according to our present knowledge, standard methodology and also to investigate the
neither the GC-FID method for the analysis behavior of degradation of lidocaine HCl with
of lidocaine HCl alone in any pharmaceutical the GC-FID method and NMR spectroscopy.
formulation nor the study of GC-FID and NMR The forced degradation studies were assessed
spectroscopy for the degradation behavior of according to the ICH guideline Q1A (R2)
lidocaine HCl under various ICH-prescribed (33). The proposed method was validated with
degradation conditions is available in the validation parameters, which are specificity,
literature. There are a few studies about the linearity, stability, analytical recovery,
stability of lidocaine HCl. These studies are sensitivity, precision and accuracy in accordance
described as follows: the relationship between with International Conference on Harmonization
pH and concentrations of antioxidants and (ICH) guidelines (42).

660
Investigation of Behavior of Forced Degradation of Lidocaine

Figure 2. GC Chromatograms of standard solutions of lidocaine HCl and IS.

Experimental chloroform (CDCl3), depending on the solubility

of the product. Chemical shifts are reported in d
Lidocaine HCl that was used as reference ppm downfield with respect to tetramethylsilane
substance and prilocaine HCl that was used as as an internal standard in 1H and 13C NMR
internal standard (IS; Figure 1 B) were purchased spectra.
from Novagenix Bio Analytical R&D Centre Photochemical experiments were performed
(Ankara, Turkey). All other chemicals were in the glass tube of a photo reactor (RPR-200).
purchased from Merck (Germany). All gases UV radiation was obtained from 24 W, RPR (The
were supplied by Havas (Ankara, Turkey). Rayonet Photochemical Chamber Reactor) that
The following pharmaceutical formulations of irradiated at 350 - 400nm (λmax = 365 nm). The
lidocaine HCl were obtained from local sources lamps were installed inside of the reactor and the
in Erzurum (Turkey): Emla® Cream (5% cream, entire system was placed in a closed box to avoid
Astra Zeneca A.S., Turkey) containing 25 mg of the disturbing effect of direct sunlight irradiation.
lidocaine HCl, 25 mg of prilocaine HCl, 55 mg
of poloxamer188 and 155 mg of poloxamer407. Chromatographic Conditions
Jetokain® Ampoule (2 mL ampoule, Adeka A.S., Separation was achieved using a HP-5 capillary
Turkey) containing about 20 mg of lidocaine HCl, column (5% - phenyl -methylpolysilocone 30 m
0.125 mg of epinephrine, 4.5 mg of sodium chloride x 0.320 mm i.d., 25-µm film thickness, USA)
and 1.0 mg of sodium metabisulfite per mL. with 2.0 μL of injector volume. The split mode
Jetmonal® Ampoule (2% 5 mL ampoule, Adeka (10:1) was used with nitrogen carrier gas and
A.S., Turkey) containing about 20 mg of lidocaine the flow rate of carrier gas was kept constant
HCl and 4.6 mg of sodium chloride per mL. during analysis at 1.6 mL/min. Hydrogen and
synthetic air were used as auxiliary gases for
Equipment the detector (FID). The injector and detector
The equipment used was: an Agilent 6890N temperatures were 260°C. The oven temperature
Network gas chromatography system provided programs were as follows: initial temperature
with an Agilent 7683 series auto sampler, a split/ of 80°C, held for 1.5 min at 210oC, ramp rate
splitless injector, flame ionization detector and of 10°C/min and final temperature of 230°C,
Agilent chemstation software. where the temperature was held for 0.5 min. The
1
H and 13C NMR spectra were recorded chromatograms under these circumstances were
on a Varian 400 spectrometer in deuterated shown in Figure 2.

661
Kadioglu Y et al. / IJPR (2013), 12 (4): 659-669

Table 1. System suitability study of method (5.0 µg/mL).

Retention time (min)a Peak areaa
7.53 22.58
SD 0.09 0.66
RSD % 1.23 2.92
a
based on six analysis, SD: standard deviation, RSD: relative standard deviation

Preparations of standard solutions Results

The standard working (SW) solutions (0.1,
0.25, 0.5, 1.0, 2.5 and 5.0 µg/mL) and quality System suitability
control (QC) solutions (0.2, 1.25 and 4.0 µg/ The system suitability tests were assessed by
mL) were prepared in methanol from stock six replicate analyses of 5.0 µg/mL concentration
solution (100 µg/mL). of lidocaine HCl. In this study, the acceptance
SW solution of prilocaine HCl (IS) was criterion was considered as ±2% deviation from
prepared at 2.0 µg/mL concentration with the percent relative standard deviation (RSD %)
methanol from the stock solution (50 µg/mL). of peak area and retention time of lidocaine HCl
(Table 1). The number of theoretical plates (N)
Procedure of pharmaceutical formulations and the height of theoretical plate [(mm), mean
Injections (RSD %)] that was expressed as the efficiency of
After the average volume for one ampoule the column were found to be 186712.6 (2.56%)
was determined, the contents of ten ampoules and 0.161 (1.56%), respectively, for injections of
for both injection samples were mixed into a six replicate.
volumetric flask. An aliquot of the solution
equivalent to 20 mg lidocaine HCl was Linearity
quantitatively transferred to a 50 mL-calibrated Linearity of method was demonstrated
measuring flask and made up to the mark with over a linear range of 0.1- 5.0 µg/mL at six
methanol. The solutions were filtered through concentrations with 2.0 µg/mL concentration of
a 0.22-µm Millipore filter. The filtrate was IS (n = 6). The calibration curve was plotted by
diluted with methanol to obtain a 0.5 µg/ the ratio of the peak areas of lidocaine HCl and
mL concentration of lidocaine HCl for both IS, versus the concentration of lidocaine HCl.
injection samples. The linear regression equation and statistical
parameters was calculated by the least squares
Cream method using Microsoft Excel® program and
An amount of the cream containing summarized in Table 2.
approximately 25 mg of lidocaine HCl was
transferred with methanol to a centrifuge Sensitivity
tube, and vortexed for 5 min. This solution The limit of detection (LOD) is the lowest
was centrifuged for 5 min at 4000 rpm. The amount of analyte in a sample that can be detected
supernatant was filtered through a 0.22-µm but not necessarily quantitated as an exact value.
Millipore filter into a volumetric flask. The The limit of quantitation (LOQ) is the lowest
filtrate was adjusted to volume with methanol to amount of analyte that can be quantitatively
obtain a 0.5 µg/mL concentration of lidocaine determined with suitable precision (42). The
HCl for the cream sample. LOD and LOQ values were determined as 3:1
About 2.0 µg/mL concentration of IS was and 10:1 of the signal/noise ratio, respectively,
added into the solutions prepared from cream and given in Table 2.
and injections. The solutions were analyzed
as described in section “Chromatographic Selectivity
Conditions”. The selectivity of an analytical method may be

662
Investigation of Behavior of Forced Degradation of Lidocaine

Table 2. Results of regression analysis of lidocaine HCl.

Parameters
Linearity (µg/mL) 0.1-50
a b
Regression Equation y = 1.726x + 0.0004
Standard Deviation of Slope (Sa) 7.52 x 10-7
Standard Deviation of Intercept (Sb) 2.96 x10-2
Correlation Coefficient 0.9992
Standard Deviation of Correlation Coefficient
4.62x10-3
Relative Residual Standard Deviation (S∆y/y, n-2)
5.88x10-2
Limit of Detection (LOD, µg/mL) 0.03
Limit of Quantitation (LOQ, µg/mL) 0.09
a
Average of six replicate determinations. b y: Peak-area ratios (lidocaine HCl/IS), x: lidocaine HCl concentration.

defined as ability to unequivocally determine the The intermediate precision was studied by
analyte in the presence of additional components analyzing the same samples on ten different
such as impurities, degradation products and days with the same procedure. The precision of
matrix (42). The method of standard addition the method was reported as the relative standard
was used to demonstrate the effective separation deviation [RSD % = (100 x standard deviation)
of lidocaine HCl in the presence of excipients / mean] and the accuracy of the method was
of pharmaceuticals and its degradation products. given with percent relative error [RE % = (found
QC samples of lidocaine HCl were spiked into concentration − known concentration) x 100 /
the solutions in equal amounts of six samples of known concentration]. The RSD % values for
20 mg (for injections) and 25 mg (for cream) of within-day and between-day precision for the
lidocaine HCl and also IS was added into each proposed method were found to be ≤1.91% (n
sample. The samples were analyzed. Peak area = 6). The RE % values for the within-day and
ratios of lidocaine HCl and IS were measured for between-day accuracy studies were found to be
quantitative determination of lidocaine HCl. Any ≤5.0% (Table 3).
interference coming from the pharmaceutical
formulations was observed. Analytical recovery
To double check the accuracy of the proposed
Precision and accuracy method, the standard addition technique was
Assay precision was determined by applied. The QC solutions of standard sample
repeatability (within-day) and intermediate were added to 0.5 µg/mL concentration of
precision (between-day). Repeatability was solutions of pharmaceutical formulations
evaluated by assaying six replicate analyses at (injections and cream) and assayed with
the three concentrations during the same day. same procedure. The analytical recovery was

Table 3. Precision and accuracy of method.

Within-day Between-day
a a
Added Found±SD Precision Accuracy Found±SD Precision Accuracy
( µg/mL ) ( µg/mL ) b
RSD% c
RE% ( µg/mL ) b
RSD% c
RE%

0.20 0.19±0.002 1.05 -5.00 0.21±0.004 1.91 5.00

1.25 1.26±0.02 1.59 0.80 1.27±0.01 0.79 1.60
4.00 3.95±0.03 0.76 1.12 4.04±0.04 0.99 1.00
a b c
SD: standard deviation RSD: relative standard deviation, RE: relative error, (n = 6).

663
Kadioglu Y et al. / IJPR (2013), 12 (4): 659-669

Table 4. Analytical recovery values by the standard addition method (n=6).

Pharmaceutical Taken Amount Added Amount Total Found Amount (µg/mL) Recovery RSD
Formulations (µg/mL) (µg/mL) (mean ±SD) (%) (%)

0.20 0.698±0.01 99.6 1.43

Jetokain® Ampoule 0.50 1.25 1.741±0.08 98.2 4.60
4.00 4.496±0.06 99.2 1.33
0.20 0.696±0.02 99.2 2.87
Jetmonal® Ampoule 0.50 1.25 1.739±0.05 97.8 2.88
4.00 4.501±0.08 100.2 1.78
0.20 0.684±0.01 96.8 1.46
0.50
Emla® Cream 1.25 1.730±0.02 96.0 1.16
4.00 4.492±0.04 98.4 0.89

calculated from the equation 1: 1

H-NMR (CDCl3, 400 MHz, ppm) δ=1.21 (t,
Analytical Recovery % = [(Ct-Ca) / Cu] x 100 J=6.9 Hz, 6H), 2.10 (s, 6H), 3.15 (q, J=6.9 Hz,
(1) 4H), 4.09 (brs, 2H), 7.26-6.91 (m, 3H).
13
C-NMR (CDCl3, 100 MHz, ppm) δ=164.9,
where Ct is total concentration of the analyte 135.4, 133.7, 128.2, 127.5, 52.5, 48.7, 18.6, 10.2.
determined, Ca is the concentration of the pure
analyte added to the formulation, and Cu is Acid-induced degradation with HCl
the concentration of the present analyte in the The solution of lidocaine HCl (0.43 mmol)
formulation. The average percent recovery was heated to reflux temperature and 0.5 mL of
values were determined approximately 97.07% concentrated (35%) hydrochloric acid solution
and 99.03% for cream and injections samples, was added slowly to this solution and stirred
respectively, indicating good accuracy of the magnetically during 24 h, 48 h and 72 h at
method. No interference from the common the same temperature. In addition, 1 M HCL
excipients was observed. The RSD % values was added into the lidocaine HCl solution and
of recovery studies were found as ranged from stirred magnetically during 24 h, 48 h and 72 h
0.89% to 4.60% (Table 4). at reflux temperature. After the stirring process,
the reaction mixture was neutralized by NaHCO3
Hydrogen peroxide-induced degradation solution and then extracted with ethyl acetate
To study hydrogen peroxide-induced (3 x 25 mL). After that, the organic layer was
degradation, the drug solution (0.43 mmol) evaporated and monitored by proton NMR
was prepared by dissolving 100 mg of powder spectroscopy.
lidocaine HCl in 50 mL methanol, and then 0.25 1
H-NMR (CDCl3, 400 MHz, ppm) δ= 1.28 (t,
mL H2O2 (35%) was added to solution. This J=7.2 Hz, 6H), 2.11 (s, 6H), 2.14 (q, J=7.2 Hz, 4H)
solution was kept at room temperature for 24 h, 3.06 (brs, NH), 3.28 (m, 2H), 6.93-7.05 (m, 3H).
48 h and 72 h in the dark brown glass containers 13
C-NMR (CDCl3, 100 MHz, ppm) δ=163.7,
and then the reaction mixture was evaporated 135.7, 135.4, 132.7, 128.2, 52.7, 49.7, 18.2, 9.4.
to completely remove the excess of hydrogen
peroxide under a nitrogen stream at room Acid-induced degradation with trifluoroacetic
temperature. The residue was monitored by acid (TFA)
proton NMR spectroscopy. A new product was The solution of lidocaine HCl (0.43 mmol)
not observed in result of reaction. However, we was heated to reflux temperature and 1 mL of
observed only a small spectral change in 1H and trifluoroacetic acid was added to this solution.
13
C-NMR spectrum. We proposed that n-oxide The reaction mixture was stirred magnetically
was obtained in reaction conditions. for 24 h and 48 h at the same temperature. After

664
Investigation of Behavior of Forced Degradation of Lidocaine

the stirring process, the mixture was evaporated, by proton NMR spectroscopy. The degradation
and the residue was purified with ethyl acetate: products were not observed.
n-hexane mixture (3:7, v/v) on silica gel column
chromatography. The final products were Photolytic degradation
determined by NMR spectroscopy. In the photolytic stability study, two solutions
were prepared in acetone. The first solution includes
2,5-dimethyl aniline 100 mg (0.215 mmol) of lidocaine HCl and the
1
H-NMR (CDCl3, 400 MHz, ppm) δ=2.10 (s, second solution was prepared by adding 5 mg of
6H), 7.25-6.92 (m, 3H), p-hydroquinone, used as a sensitizer solution, and
13
C-NMR (CDCl3, 100 MHz, ppm) δ= 135.2, 100 mg of lidocaine HCl into 25 mL of acetone.
133.2, 128.3, 127.8, 18.3. To start the experiments, the lamps of the photo
reactor (RPR) were turned on and warmed up for
2-(diethylamino)acetic acid about 10 min. After that, the prepared solutions
1
H-NMR (CDCl3, 400 MHz, ppm) δ=1.25 (t, were placed into the reactor’s glass tube and then
J=6.8 Hz, 6H), 3.23 (brq, J= 6.8 Hz, 4H), 4.17 the photochemical reaction at room temperature
(brs, 2H) 10.01 (brs, 1H). 13C-NMR (CDCl3, 100 was started and exposed to light (365 nm) during
MHz, ppm) δ=163.5, 52.0, 48.9, 9.3 24 h, 48 h and 72 h. Under illumination, the
reaction temperature was kept at 20 - 25°C. After
Base-induced degradation studies this process, the reaction solution was evaporated
In the study in alkaline condition, the solution and the residue was purified on the silica gel
of lidocaine HCl (0.43 mmol) was heated to column chromatograph. The reaction mixture was
reflux temperature and 2.5 mL of 1.0 M NaOH monitored by proton NMR spectroscopy. Any
solution was added slowly to this solution. The degradation products were assessed.
reaction mixture was stirred magnetically for
24 h, 48 h and 72 h at the same temperature. Application of the method for analysis of
After the stirring process, the mixture was pharmaceutical formulations
neutralized by 1.0 M HCl solution and then The proposed method was evaluated in
diluted. The reaction mixture was extracted with the assay of commercially available a cream
ethyl acetate (3 x 25 mL), and the organic layer containing 25 mg of lidocaine HCl and two
was evaporated. The residue was monitored by brands of injections containing 20 mg of
proton NMR spectroscopy. The degradation lidocaine HCl. Assessment was created using
products were not observed. the calibration curve method. Any important
difference between the slopes of the calibration
Dry heat degradation curves of pharmaceutical formulation and
In order to observe the effect of temperature standard solutions was observed. The accurately
on degradation of lidocaine HCl, 50 mg (0.215 weighted amounts of ampoules and cream
mmol) powder lidocaine HCl was exposed to including 20 mg and 25 mg of lidocaine HCl,
dry heat at 120°C and kept for 24 h, 48 h and respectively, were determined (n = 6).
72 h at same temperature. After that, the residue The results obtained are satisfactorily accurate
was cooled to room temperature and monitored and precise, as indicated by the excellent %

Table 5. Determination of lidocaine HCl in pharmaceutical formulations.

Pharmaceutical
Label Claim a
Mean±SD Recovery (%) RSD (%)
Formulations

Jetokain ® ampoule 20 mg per mL 19.98±1.21 99.9 6.06

Jetmonal ® ampoule 20 mg per mL 19.96±1.12 99.8 5.61
Emla® cream 25 mg 20.01±0.99 100.1 4.95
aAverage of six replicate determinations.

665
Kadioglu Y et al. / IJPR (2013), 12 (4): 659-669

Figure 3. GC Chromatograms: A: the standard solution of

lidocaine HCl; B: degradation with HCl for 72 h; C: degradation Figure 4. Proton NMR spectrums of lidocaine HCl and
with H2O2 for 72 h; D: degradation with trifluoroacetic acid degradation with H2O2 and trifluoroacetic acid (TFA).
(TFA) for 72 h.

recovery and SD <1.21 (Table 5). Experiments indication of the stability-indicating properties
showed that there was no interference from the of lidocaine HCl, and the forced degradation
additions and excipients. In the determination studies were performed under various stress
repeated six times, mean recovery for all conditions. The suitability of the proposed
formulations obtained approximately 99.93%, method as a stability- indicating method was
with an RSD% <5.54%. supported by NMR studies. After the forced
degradation process, final products were diluted
Discussion with methanol to obtain 4 µg/mL solutions and
injected into the GC system. The chromatograms
In the pharmaceutical industry, the obtained are presented in Figure 3. In addition,
disciplines primarily involved with stability the degradation reaction mixture was monitored
are pharmaceutical analysis and product by NMR spectroscopy (Figure 4).
development. The requirement of the According to the degradation results, it was
establishment of a stability-indicating assay observed that lidocaine HCl has a very stable
has become more clearly mandated with the skeleton structure. In acidic conditions, when
International Conference on Harmonization lidocaine HCl was treated with HCl acid, it was
(ICH) guidelines (33). The guidelines explicitly observed that secondary amine salt of lidocaine
require conduct of forced degradation studies was formed to yield 100%. In proton NMR
under a variety of conditions, like pH (acid- spectroscopy, resonance structure was produced
base), photolytic, oxidation and dry heat (33, 43). as multiple of doublet in 3.28 ppm. Since, each
Understanding drug degradation in formulated of CH2 protons has the site of different spaces.
product is important in pharmaceutical It was observed that proton NMR signals
development as drug stability and degradation obtained were the same with molecules of
products could have significant impacts on similar nature in the literature (44). When
formulation development, analytical method trifluoroacetic acid (TFA) was used as the
development, package development, storage acidic media, as different from International
conditions, shelf-life determination, and safety Conference on Harmonization (ICH)
and toxicology concerns (43). Consequently, guidelines, decomposition products were
we aimed to designate the GC-FID method for observed (Scheme 1).

666
Investigation of Behavior of Forced Degradation of Lidocaine

Scheme 1

Oxidation of lidocaine HCl with H2O2 and analysis of degradation products during the
was done at room temperature, and a small stability studies.
change in 1H-NMR spectrum of lidocaine was
observed as a result of the oxidation reaction. References
Hence, we proposed that nitrogen groups in the
structure of lidocaine HCl were oxidized with (1) Cawson RA and Curson DR. The hazards of dental
H2O2 as n-oxide. In alkali conditions, skeleton local anesthetics in children. Br. Dent. J. (1983) 154:
structure of lidocaine HCl was protected. After 253-256.
(2) Li YM, Wingrove DE, Too HP, Marnerakis M, Stimson
dry heat and photolytic studies we determined ER, Strichartz GR and Maggio JE. Local anesthetics
that lidocaine HCl was very stable against heat inhibit substance P binding and evoked increases in
conditions (120 oC) and light (365 nm) over 72 h. intracellular Ca2+. Anesthesiology (1995) 82: 166-173.
(3) Rishiraj B, Epstein JB, Fine D, Nabi S and Wade
Conclusions NK. Permanent vision loss in one eye following
administration of local anesthesia for a dental
extraction. Int. J. Oral Max. Surg. (2005) 34: 220-223.
A rapid and sensitive gas chromatography (4) Zivanovic LJ, Zivanov-Stakic D and Radulovic
method with flame ionization detection (GC- D. Determination of lidocaine in pharmaceutical
FID) was developed and validated for the preparations using thin-layer chromatographic
determination of lidocaine HCl in pharmaceutical densitometry. J. Pharm. Biomed. Anal. (1988) 6: 809-
formulations in the form of cream and injections. 812.
(5) Oliveira RTS, Salazar-Banda GR, Valdir S, Ferreira
The proposed method has several advantages, VS, Oliveira SC and Avaca LA. Electroanalytical
which are high specificity, good accuracy and determination of lidocaine in pharmaceutical
precision values, short chromatographic run time preparations using boron-doped diamond electrodes.
(8.5 min) and high analytical recovery value. Electroanalysis (2007) 19: 1189-1194.
In forced degradation studies, many forms of (6) Nerin C, Garnica A and Cacho J. Indirect determination
of lidocaine by atomic absorption spectrophotometry.
degradation and fragmentation were not observed
Anal. Lett. (1991) 24: 1847-1859.
in destruction reactions. It was found that the (7) El-Ries MA, Abou Attia FM, Abdel-Gawad FM and
structure of the lidocaine HCl molecule was very Abu El-Wafa SM. Preparation and characterization
stable in alkaline, acidic, oxidation, dry heat and of Cu(II)-lignocaine complex. Indirect estimation of
photolytic media. Furthermore, trifluoroacetic lignocaine in pharmaceutical preparations by atomic
acid (TFA) was used as the acidic media. In this absorption spectroscopy (AAS). J. Pharm. Biomed.
Anal. (1994) 12: 1209-1213.
media, the decomposition products occurred. The (8) Ismaiel SA, Yassa DA and Gad-El-Rub LN.
developed GC-FID method, together with NMR Spectrophotometric determination of lidocaine in
spectroscopy, for determination of lidocaine HCl some pharmaceutical preparations using bromocresol
in pharmaceutical formulations can be used for green. Pharmazie (1975) 30: 408-410.
both routine analysis in quality control studies (9) Rizk MS, Issa YM, Shoukry AF and Atia EM.

667
Kadioglu Y et al. / IJPR (2013), 12 (4): 659-669

Spectrophotometric determination of lignocaine in anesthetic formulations. J. Assoc. Off. Anal. Chem.

pure form and pharmaceutical preparations. Anal. Lett. (1986) 69: 618-620.
(1997) 30: 2743-2753. (23) Ohshima T, and Takayasu T. Simultaneous
(10) Escuder-Gilabert L, Sagrado S, Villanueva- determination of local anesthetics including ester-
Camañas RM and Medina-Hernandez MJ. Analysis type anesthetics in human plasma and urine by gas
of pharmaceutical preparations containing local chromatography-mass spectrometry with solid-phase
anesthetics by micellar liquid chromatography and extraction. J. Chromatogr. B (1999) 726: 185-194.
spectrophotometric detection. Chromatographia (24) Keenaghan JB. The determination of lidocaine and
(1999) 49: 85-90. prilocaine in whole blood by gas chromatography.
(11) Mirjana M, Darko I and Andelija M. Simultaneous Anesthesiology (1968) 29: 110-112.
determination of compounds in Septalen(R) pellets (25) Asada A. Gas chromatographic determination of local
by derivative spectrophotometry. J. Serb. Chem. Soc. anesthetics in blood. Osaka City Med. J. (1979) 25:
(2000) 65: 339-344. 91-122.
(12) Nemcova I, Rychlovsky P, Tomankova V and Zivanovic (26) Prat M and Bruguerolle B. Rapid simultaneous gas-
Lj. Extraction spectrophotometric determination of liquid chromatographic determination of five local
lidocaine using flow injection analysis. Anal. Lett. anesthetic drugs in serum. Clin. Chem. (1986) 32:
(2001) 34: 2457-2464. 2098- 2100.
(13) Parissi-Poulou M and Panderi I. Determination of (27) Björk M, Pettersson KJ and Osterlöf G. Capillary
hyoscine n-butylbromide, lidocaine hydrochloride, gas chromatographic method for simultaneous
and paracetamol in injection forms using solid-phase determination of local anaesthetics in plasma samples.
extraction, high performance liquid chromatography J. Chromatogr. (1990) 533: 229-234.
and UV-VIS spectrophotometry. J. Liq. Chromatogr. (28) Baniceru M, Croitoru O and Popescu SM.
Relat. Technol. (1999) 22: 1055-1068. Determination of some local anesthetics in human
(14) Liawruangrath S, Liawruangrath B and Pibool P. serum by gas chromatography with solid-phase
Simultaneous determination of tolperisone and extraction. J. Pharm. Biomed. Anal. (2004) 35: 593-
lidocaine by high performance liquid chromatography. 598.
J. Pharm. Biomed. Anal. (2001) 26: 865-872. (29) Tarli P, Benocci S and Neri P. Gas chromatographic
(15) Youngvises N, Liawruangrath B and Liawruangrath S. determination of nicotinamide and lidocaine in
Simultaneous micellar LC determination of lidocaine pharmaceutical preparations. Boll. Chim. Farm. (1969)
and tolperisone. J. Pharm. Biomed. Anal. (2003) 31: 108: 554-559.
629-638. (30) Edhorn GA. Determination of lidocaine in whole
(16) Fijałek Z, Baczyήski E, Piwoήska A and Warowna- blood by gas chromatography. Can. Anaesth. Soc. J.
Grzeskiewicz M. Determination of local anaesthetics (1971) 18: 189-198.
and their impurities in pharmaceutical preparations (31) Kruczek ME. A rapid gas-liquid chromatographic
using HPLC method with amperometric detection. J. determination of serum lidocaine using a nitrogen-
Pharm. Biomed. Anal. (2005) 37: 913-918. phosphorus specific detector. J. Pharmacol. Methods
(17) Mohammad MAA. LC determination of lidocaine (1981) 5: 137-141.
and prilocaine containing potential risky impurities (32) Koster EHM, Hofman NSK and de Jong GJ. Direct
and application to pharmaceuticals. Chromatographia solid-phase microextraction combined with gas
(2009) 70: 563-568. and liquid chromatography for the determination of
(18) Jancic-Stojanoviċ B, Malenoviċ A, Markoviċ S, lidocaine in human urine. Chromatographia (1998)
Ivanoviċ D and Medenica M. Optimization and 47: 678-684.
validation of a RP-HPLC method for analysis of (33) Stability Testing of New Drugs Substances and
hydrocortisone acetate and lidocaine in suppositories. Products. ICH Topic Q1A (R2), CPMP/ICH/2736/99,
J. Assoc. Off. Anal. Chem. Int. (2010) 93: 102-107. European Medicines Agency, London, UK, (2003).
(19) Shaalan RA and Belal TS. HPLC-DAD stability (34) Hondrum SO and Ezell JH. The relationship
indicating determination of nitrofurazone and lidocaine between pH and concentrations of antioxidants and
hydrochloride in their combined topical dosage form. vasoconstrictors in local anesthetic solutions. Anesth.
J. Chromatogr. Sci. (2010) 48: 647-653. Prog. (1996) 43: 85-91.
(20) Candioti LV, Robles JC, Mantovani VE and Goicoechea (35) Tuleu C, Allam J, Gill H and Yentis SM. Short term
HC. Multiple response optimization applied to the stability of pH-adjusted lidocaine-adrenaline epidural
development of a capillary electophoretic method for solution used for emergency caesarean section. Int. J.
pharmaceutical analysis. Talanta (2006) 69: 140-147. Obstet. Anesth. (2008) 17: 118-122.
(21) Ionescu MS, Abrutis AA, Radulescu N, Baiulescu (36) Robinson J, Fernando R, Sun Wai WYS and Reynolds
GE and Coşofret VV. Lidocaine-selective membrane F. Chemical stability of bupivacaine, lidocaine and
electrodes and their applications in pharmaceutical epinephrine in pH-adjusted solutions. Anaesthesia
analysis. Analyst (1985) 110: 929-931. (2000) 55: 853-858.
(22) Hassan SS and Ahmed MA. New liquid membrane (37) Gebauer MG, McClure AF and Vlahakis TL. Stability
electrode for determination of lidocaine in local indicating HPLC method for the estimation of

668
Investigation of Behavior of Forced Degradation of Lidocaine

oxycodone and lidocaine in rectal gel. Int. J. Pharm. and lidocaine hydrochloride in their combined oral gel
(2001) 223: 49-54. dosage form. J. Chromatogr. Sci. (2012) 50:401-409.
(38) Di Stasi SM, Giannantoni A, Navarra P, Massoud R, (41) Katz J. Stability of local anesthetics after autoclaving:
Zavaqlia D, Bertucci P, Vespasiani G and Stephen determined by gas chromatography. Anesthesiology
RL. The stability of lidocaine and epinephrine (1966) 27: 835-837.
solutions exposed to electric current and comparative (42) ICH Harmonised Tripartite Guideline, Validation of
administration rates of the two drugs into pig bladder Analytical Procedures: Text and Methodology Q2
wall. Urol. Res. (2003) 31: 169-176. (R1), Geneva, (2005).
(39) Waraszkiewicz SM, Milano EA and DiRubio (43) Mollica JA, Ahuja S, and Cohen J. Stability of
R. Stability-indicating high-performance liquid pharmaceuticals. J. Pharm. Sci. (1978) 67: 443-465.
chromatographic analysis of lidocaine hydrochloride (44) Hanna GM. H-1-NMR studies of drugs with chiral
and lidocaine hydrochloride with epinephrine solvating agent: The direct enantiomeric purity
injectable solutions. J. Pharm. Sci. (1981) 70: 1215- determination of the chiral anesthetic, prilocaine.
1218. Enantiomer (2000) 5: 303-312.
(40) Belal TS and Haggag RS. Gradient HPLC-DAD This article is available online at http://www.ijpr.ir
stability indicating determination of miconazole nitrate

669