Validated HPLC and stability-indicating densitometric chromatographic methods for simultaneous determination of camylofin dihydrochloride and paracetamol in their binary mixture

Article abstract of DOI:10.1007/s13738-020-01953-8

Two accurate, sensitive, precise and selective HPLC and stability-indicating TLC methods were developed for the simultaneous determination of camylofin-2HCl and paracetamol. Forced acid, alkali and oxidative degradation of camylofin-2HCl?were tried where complete degradation was achieved using 5?N HCl. HPLC method was developed to determine the mixture of the two drugs using Zorbax NH₂ column and a mobile phase of 0.5percent triethylamine and pH 3.0 adjusted with 0.1percent phosphoric acid and methanol (70:30 v/v) over concentration ranges of 3–90 and 10–95?μg/mL for camylofin-2HCl and paracetamol, respectively.TLC method was used for the separation of camylofin from its acid degradate and paracetamol using chloroform–methanol–acetone–conc. ammonia (8:2:2:0.1, by volume) as developing system and band scanning at 254 nm over concentration ranges of 5–40?μg/band for camylofin-2HCl and 0.1–0.5?μg/band for paracetamol. The validation of two methods was carried out according to ICH guideline. Accuracy ranged between 98.47 and 100.67percent for the two methods with acceptable precision RSDpercent ranging between 0.66 and 1.47percent.

Full text of DOI:10.1007/s13738-020-01953-8

Journal of the Iranian Chemical Society
https://doi.org/10.1007/s13738-020-01953-8
ORIGINAL PAPER
Validated HPLC and stability‑indicating densitometric
chromatographic methods for simultaneous determination
of camyloꢀn dihydrochloride and paracetamol in their binary mixture
Sawsan A. Abdel Razeq · Israa A. Khalil1 · Samah A. Mohammd2
1
Received: 7 January 2020 / Accepted: 16 May 2020
©
Iranian Chemical Society 2020
Abstract
Two accurate, sensitive, precise and selective HPLC and stability-indicating TLC methods were developed for the simul-
taneous determination of camyloꢀn-2HCl and paracetamol. Forced acid, alkali and oxidative degradation of camyloꢀn-
2
HCl were tried where complete degradation was achieved using 5 N HCl. HPLC method was developed to determine the
mixture of the two drugs using Zorbax NH column and a mobile phase of 0.5% triethylamine and pH 3.0 adjusted with
2
0
.1% phosphoric acid and methanol (70:30 v/v) over concentration ranges of 3–90 and 10–95 µg/mL for camyloꢀn-2HCl
and paracetamol, respectively.TLC method was used for the separation of camyloꢀn from its acid degradate and paracetamol
using chloroform–methanol–acetone–conc. ammonia (8:2:2:0.1, by volume) as developing system and band scanning at 254
nm over concentration ranges of 5–40 µg/band for camyloꢀn-2HCl and 0.1–0.5 µg/band for paracetamol. The validation of
two methods was carried out according to ICH guideline. Accuracy ranged between 98.47 and 100.67% for the two methods
with acceptable precision RSD% ranging between 0.66 and 1.47%.
Keywords Camyloꢀn · Paracetamol · Acidic degradate · HPLC · TLC densitometry
Introduction
[22–25], GC [26–29], ꢁuorometry [30–32] and electrochem-
istry [33–36].
Camyloꢀn-2 HCl (3-methyl butyl 2-(2-diethyl amino ethyl
amino)-2-phenyl acetate hydrochloride) is a part of anticho-
linergic, spasmolytic and gastrointestinal sedative group [1,
Literature review of the mixture analysis revealed that
there are only two reported HPLC methods [37, 38] and one
spectrophotometric method [39] which were developed for
the determination of both drugs simultaneously. But all the
reported methods lack a sensitivity, and no reported TLC
densitometric method for the simultaneous determination of
camyloꢀn-2HCl and paracetamol was developed up till now.
The novelty of this work is the ꢀrst development of TLC
densitometric method for the determination of paracetamol
and camyloꢀn-2HCl in the presence of its acidic degradate.
The value of this work over all the published methods is the
complete degradation of camyloꢀn and the identiꢀcation of
2
]. It is found in market as single or combined formulation
with paracetamol, nimesulide, diclofenac and analgin. The
literature revealed many analytical techniques for the quan-
tiꢀcation of camyloꢀn-2HCl such as spectrophotometry [3,
4
], HPLC [5–9] TLC densitometry [6, 7, 10–12] and GC
[
13, 14].
Paracetamol (N-acetyl-para-amino-phenol) has antipy-
retic and analgesic action and weak anti-inꢁammatory eꢂect
1]. Many techniques have been used for its determination,
including spectrophotometry [15–17], HPLC [18–21] TLC
[
1
degradate by IR and HNMR with the suggestion of degra-
dation pathway which matched that reported uncompleted
degradation [8]. Also the proposed HPLC method has an
advantage of high sensitivity over the two reported ones [37,
*
Israa A. Khalil
p_isr@yahoo.com
3
8] (Fig. 1).
1
2
Analytical Chemistry Department, Faculty of Pharmacy
(
Girls), Al-Azhar University, Cairo, Egypt
National Organization for Drug Control and Research
NODCAR), Giza, Egypt
(
Vol.:(0
1
123456789)
3

Journal of the Iranian Chemical Society
Working standard solutions for both drugs (100.00 µg/
mL) for HPLC and (5.0 mg/mL) camyloꢀn-2-HCl, (0.05 mg/
mL) paracetamol for TLC were prepared from the stock
solutions by dilution with water for HPLC and by methanol
for TLC method.
Degraded sample
Acid and basic degradation
Fig. 1 Chemical structure of a camyloꢀn and b paracetamol
Acidic and basic degradates of camyloꢀn-2HCl were pre-
pared by reꢁuxing 0.5 g pure drug with either 100 mL 5 N
HCl or 5 N NaOH for about 6 h. The solution was neutral-
ized with either 5 N NaOH or 5 N HCl to pH about 7 and
evaporated till dryness under vacuum. Residue was extracted
three times, each with 30 mL methanol, and then ꢀltered
into a 100-mL volumetric ꢁask followed by dilution with
the same solvent. Portion of the prepared methanolic degra-
date was then evaporated to dryness, and the structure of the
Experimental
Instruments
•
HPLC (Agilent 1100 series, Waldbronn, Germany) consists
of an Agilent pump, equipped with autosampler and vari-
able wavelength detector.
1
degradate was conꢀrmed by IR and H NMR.
•
•
•
•
An Agilent Zorbax NH column (250×4.6 mm, 5 μm par-
2
ticle).
Oxidative degradation
TLC scanner with WINCATS software (Camag, Switzer-
land).
The above steps for acidic and basic degradation were fol-
lowed using 30% H O for oxidative degradation without
TLC plates precoated with Silica Gel 60 F254, 10×20 cm
2
2
(Merck, Germany).
reꢁuxing.
An automatic sample applicator provided with 100 µL
syringe.
Chromatographic conditions
HPLC method
•
•
Five-digit electronic balance (Radwag, Germany).
Sonicator (Crest, USA).
Materials and reagents
Chromatographic separation was performed on Zorbax NH
2
column (250×4.6 mm, 5 μm particle) using water contain-
ing 0.5% triethylamine, and pH was adjusted to 3 by 0.1%
phosphoric acid/methanol (70:30 v/v) as mobile phase
pumped with ꢁow rate of 1 mL/min and UV detection at
Pure camylofin-2HCl and paracetamol were supplied by
Kahira Pharmaceutical Co. (Cairo, Egypt) and Chemical
Industries Co, (Cairo, Egypt) with purity of 99.64% and
®
9
9.85% [38]. Anafortan tablet which contains 25 mg of
2
20 nm.
camyloꢀn-2HCl and 300 mg paracetamol and is the product of
Abbot, (India) was purchased from a local pharmacy in India.
All the reagents were of HPLC grade; methanol and tri-
ethylamine were supplied from Fisher scientiꢀc, USA, and
chloroform and acetone were supplied from Scharlau Chemie,
Spain, while orthophosphoric acid and ammonia were sup-
plied from Sigma, USA. Water for HPLC was prepared by
double distillation and then ꢀltration using 0.2 μm membrane
TLC method
Camylofin-2HCl, its acidic degradate and paracetamol
solutions were applied to a Silica Gel 60 F254 plates
(
20× 10 cm) using automatic applicator and a developing
system of chloroform/methanol/acetone/conc. ammonia
8:2:2:0.1, by volume). The plates were scanned at 254 nm
under the following conditions;
(
ꢀlter.
Standard solutions
•
•
•
•
•
Silt dimensions: 6.0×0.2 mm.
Scanning speed: 20 mm/s.
Stock solutions of camyloꢀn-2HCl (10.00 mg/mL) and par-
acetamol (1.00 mg/mL) were prepared in water for HPLC
method and in methanol for TLC method.
Data resolution: 100 μm/step.
Measurement mode: absorption.
Result output: chromatogram and integrated peak area.
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Journal of the Iranian Chemical Society
Validation
An amount of powdered tablet equivalent to 25 mg of
camyloꢀn-2HCl and 300 mg of paracetamol was transferred
into a 100-mL volumetric ꢁask and dissolved in 70 ml of
water and sonicated for 15 min, and the volume was com-
pleted with water and ꢀltered. The ꢀltrate was labeled to
contain 250 μg/mL of camyloꢀn-2HCl and 3 mg/mL of
paracetamol.
(
a) Linearity and calibration curves
HPLC methodHundred microliters from solutions
containing 3.00–90.00 µg/mL or 10.00–95.00 µg/mL of
camyloꢀn or paracetamol was injected on the column and
chromatographed under the previously described conditions
under “Chromatographic conditions.”
TLC method
Camylofn-2HCl An amount of ꢀne powder equivalent
to 50 mg camyloꢀn-2HCl and 600 mg of paracetamol was
transferred into a 50-mL volumetric ꢁask, dissolved in 30 ml
of water and sonicated for 15 min, and the volume was com-
pleted with methanol and ꢀlter. The ꢀltrate was labeled to
contain 1.00 mg/mL of camyloꢀn-2HCl and 12.00 mg/mL
of paracetamol
TLC method Aliquots of 2.5–20 mg from camyloꢀn-
2
0
HCL working standard solution (5.0 mg/mL) or
.05–0.25 mg from paracetamol working standard solution
(
0.05 mg/mL) were transferred into two separate sets of
1
0-mL volumetric ꢁasks, and the volume was completed to
the mark with methanol. Twenty microliters of each solu-
tion was applied to Silica Gel 60 F254 plates. The procedure
under “Chromatographic conditions” was followed.
The calibration curves were constructed by plotting the
peak area against the corresponding drug concentration.
Regression equations were computed.
Paracetamol An amount of ꢀne powder equivalent to
50 mg of paracetamol was transferred into 100-mL volu-
metric ꢁask, dissolved in 30 ml of water and sonicated
for 15 min, and the volume was completed with methanol
and ꢀlter. Filtrate was labeled to contain 0.50 mg/mL of
paracetamol.
The procedure of “linearity” for each method was fol-
lowed, and the concentration of each drug was calculated
from its corresponding regression equation.
(
b) Accuracy
The accuracy of the suggested methods was determined
by the standard addition technique in which known concen-
trations of standard camyloꢀn and paracetamol (3,20, 85 μg/
mL and 10, 20, 30 μg/mL and 5, 10, 30 μg/band and 0.1,0.2,
Results and discussion
0
.3 μg/band) drugs for HPLC and TLC methods, respec-
Literature review revealed that no stability densitomet-
ric TLC method was reported for the determination of
camyloꢀn-2HCl and paracetamol mixture. Only two HPLC
methods [37, 38] were reported for the determination of
tively, were added to ꢀxed amount of pharmaceutical formu-
lation. The concentrations of the added standard were then
calculated from their corresponding regression equation.
1
mixture without HNMR or IR identiꢀcation of degradate.
(
c) Precision
This work introduces the ꢀrst preparation, separation and
identiꢀcation of degradate of camyloꢀn with a stability indi-
cating TLC method for determination of camyloꢀn-2HCl
which is also simultaneous for its determination with par-
acetamol. Both drugs were also determined simultaneously
with a new more simple and sensitive HPLC method than
the reported methods.
Triplicate analyses at three concentration levels of 5,
5
2
0 and 90 μg/mL and 5, 30 and 40 µg/band for camyloꢀn-
HCl and 10, 50 and 95 μg/mL and 0.1, 0.3 and 0.5 µg/band
for paracetamol were performed in the same day to study
the intraday precision of HPLC and TLC methods, respec-
tively. The interday precision was conꢀrmed by repeating
the analysis in three successive days using the same drug
concentration levels.
Forced degradation of camyloꢀn‑2HCl
Degradation of camyloꢀn-2HCl was performed by reꢁux-
ing the drug using diꢂerent media with diꢂerent normalities
and for diꢂerent time intervals: 0.1 N HCl and NaOH for
10 h, 1 N HCl and NaOH for 8 h, 3 N HCl and NaOH for
8 h, 5 N HCl and NaOH for 6 h and 10%, 20%, 30% H O
(
d) Selectivity
Selectivity was evaluated by simultaneous determination
of camyloꢀn and paracetamol for HPLC and in the presence
of camyloꢀn acid degradate for TLC method.
2
2
for 24 h. Acidic, basic and oxidative degradates were then
treated as under “degraded samples.” Complete degradation
was attained upon reꢁuxing the drug with 5 N HCl for about
(
e) Application to pharmaceutical formulations
6
h. TLC separation of the degradate was attained after test-
HPLC method
ing diꢂerent developing systems for good separation, and
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Journal of the Iranian Chemical Society
the most suitable system was chloroform/methanol/acetone/
conc. ammonia (8: 2:2: 0.1, by volume). Upon calculating of
Rf of degradate and the intact drug, acidic, basic and oxida-
tive degradates have the same Rf values. Thus, the acidic
degradate is subsequently used for the stability-indicating
analysis of the drug as the alkaline and oxidative degrada-
tion is not complete. The structure of acidic degradates was
carbon” (1) is shifted to 4 ppm in the degradate spec-
trum. The spectrum of camyloꢀn-2HCl shows signals at
3.5, 1.4, 1.2 and 0.73 ppm for H of two methylene groups
located between 2 amino groups (2), H of two methylene
of tertiary amines (3), H of methyl groups (4) and H of
(O–CH –CH ) (5), respectively, which completely disap-
2
2
1
pear in the HMNR spectrum of the degradate.
1
1
conꢀrmed by IR and HNMR (Figs. 2a, b, 3a, b, 4a, b).
HMNR spectrum of camylofin-2HCl shows a signal
1
Comparing the HNMR spectra of intact camyloꢀn-
at 3.4 ppm, which completely disappears in D O spec-
2
2
HCl and its acidic degradate shows that there is a signal
trum, which represents H for secondary amino group
at 7.3 for “H of the aromatic ring” which is also present
in both spectra. A signal at 5.2 ppm for “H of benzylic
(6), while at degradate, the spectrum shows a signal at
1
Fig. 2 a HNMR D O spectrum
2
1
of intact camyloꢀn, b HNMR
spectrum of intact camyloꢀn
1
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Journal of the Iranian Chemical Society
1
Fig. 3 a HNMR D O spectrum
2
1
of degradate, b HNMR of
degradate
1
.2 ppm which completely disappears in D O spectrum
Method development
2
which represents H of NH (7).
2
−
1
IR spectrum of degradate shows peak at 1581 cm ,
which represents carbonyl group of carboxylic acid
formed after breakage of ester bond and peak at
(a) HPLC method
−
1
3
436 cm for the formed primary amine (Fig. 3a, b).
Accordingly, a degradation pathway was suggested as
The main challenge in the development of HPLC method
was the diꢂerence in the solubility of both drugs; camyloꢀn
is freely soluble in water, while paracetamol is not. The
mobile phase should contain an appropriate amount of water
shown in Fig. 5, which is consistent with the published
finding [8].
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Journal of the Iranian Chemical Society
Fig. 4 a IR spectrum of intact camyloꢀn, b IR spectrum of degradation product
Fig. 5 Suggested degradation
pathway camyloꢀn-2HCl
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Journal of the Iranian Chemical Society
to ensure separation and to decrease retention time. Diꢂerent
mobile phase systems were tried for separation of two drugs
with diꢂerent organic ratios such as acetonitrile/methanol,
acetonitrile/phosphate buꢂer, acetonitrile/methanol/water,
acetonitrile/phosphate buꢂer, methanol/water, and the best
separation was obtained by using a mixture of water con-
taining 0.5% triethylamine with pH of 3.0 adjusted by 0.1%
phosphoric acid/methanol (70:30 v/v). Good resolution and
acceptable symmetric peaks were achieved by using Zor-
HPLC methods [37, 38], while that of paracetamol is longer
than the reported value in Ref. [37] and shorter than that
reported in Ref. [38].
(b) TLC method
Diꢂerence in the aꢃnity of the two drugs to TLC mobile
phases causes a diꢃculty in their separation by TLC. That
is why there is no published method developed for the
mixture separation till now. In this work, many developing
systems were tested for the separation of three compounds
(Table 1). First, a mobile phase composed of acetone/meth-
anol/ammonia (6:4:0.1, by volume) was tried; both drugs
moved with eluting system but there was over-peak tailing.
To increase unipolarity of the system, Systems IV, VI, VII,
VII were tried but the problem of tailing was still present.
Complete separation of the three compounds with suitable
bax—NH column (250 mm×4.6 mm, 5 μm particle) after
2
testing a variety of column types. The detection at 220 nm
was chosen to increase the sensitivity of the measurement.
Also the best resolution between peaks was obtained by
pumping the mobile phase with a ꢁow rate of 1.0 mL/min.
The retention times of two peaks were 2.047 and 3.015 for
camyloꢀn-2HCl and paracetamol, respectively (Fig. 6).
RT of camyloꢀn is shorter than that reported in previously
Fig. 6 HPLC chromatogram
of mixture of camyloꢀn-2HCl
(
(
90 µg/mL) and paracetamol
10 µg/mL)
Table 1 Diꢂerent development
systems tested for the separation
of camyloꢀn-2HCl, its acidic
degradate and paracetamol
Mobile phase component
Ratio of component, by volume
I
II
III
IV
V
VI
VII
VIII
IX
Acetone/glacial acetic acid
Acetone/methanol/ammonia
Acetone/methanol/acetic
5:5
8:2:0.1, 6:4:0.1
6.5:3.5:2.5
5:2,6:2,7:1.5
8:4:0.2, 5:1:0.1,7:1.5:0.1
10:2:0.1
5:1
1:2:2
Hexane/methanol
Hexane/methanol/ammonia
Hexane/methanol/glacial acetic acid
Hexane/ethyl acetate
Methanol/ethyl acetate/toluene
hexane/propanol
8:1.5
X
XI
XII
XIII
XIV
XV
XVI
Acetone/ethyl acetate
6:2
6:2:2
Acetone/ethyl acetate/methanol
Chloroform/methanol/ammonia
Chloroform/methanol/ammonia/hexane.
Chloroform/methanol/ammonia/toluene
Chloroform/acetone/ammonia
Chloroform/methanol/acetone/ammonia
6:2:0.1,6:4:0.1, 8:4:0.1
8:4:0.1:1
8:4:0.1:1
8:2:0.1
8:2:2:0.1
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Journal of the Iranian Chemical Society
Fig. 7 Densitometric chromato-
gram of mixture of camyloꢀn-
2
HCl (20 µg/band), the acid
degradate of camyloꢀn (20 µg/
band) and paracetamol (0.1 µg/
band) at 254 nm
peak characteristics was achieved using a mobile phase of
chloroform/methanol/acetone/conc. ammonia (8:2:2:0.1,
by volume). The Rf values were 0.84, 0.13 and 0.72 for
camyloꢀn-2HCl, its degradate and paracetamol, respectively,
by detection at 254 nm (Fig. 7). TLC method was considered
stability-indicating method with respect to camyloꢀn-2HCl.
Method validation
The two methods were validated according to ICH guideline
[42].
(a) Linearity
System suitability
Under the above-optimized experimental conditions,
linear relationship was found between the peak areas and
the corresponding drug concentration over the ranges
of 3.0–90.0 and 10.0–95.0 µg/mL for HPLC method
and 5.0–40.0, 0.10–0.50 µg/band for TLC method for
The results are shown in Tables 2, 3 indicating that both
HPLC and TLC systems are suitable.
2
camyloꢀn-2HCl and paracetamol, respectively. The high r
values (0.9993–0.9999) indicate good linearities. Regres-
sion parameters are summarized in Table 4. The proposed
method is more sensitive than the two reported ones,
which measured camyloꢀn in the concentration ranges of
20.0–80.0 μg/mL [37] and 25–75 mg/mL [38].
Table 2 System suitability data for HPLC for the determination of
camyloꢀn-2HCl and paracetamol
Parameters
Obtained value
Reference value [40]
Camyloꢀn Paracetamol
Rt=2.047 Rt=3.015
(
b) Accuracy and precision
Capacity factor
0.9
1.7
0.5–10 is acceptable
(
K’)
Accuracy was found to be 100.67% and 100.59% for
Selectivity factor
α)
Resolution factor
R)
1.47
6.13
1.704
2283
>1
camyloꢀn-2HCl and 98.47% and 99.08% for paracetamol
for HPLC and TLC methods, respectively. The results are
summarized in Table 4.
(
R>1.5
(
Tailing factor (T)
1.872
6942
T=1 for typical
peaks
(c) Selectivity
Number of plates
The higher the value,
the more eꢃcient
the column
(
N)
HPLC selectivity was determined by analyzing labora-
tory-prepared mixtures of camyloꢀn-2HCl and paracetamol
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Journal of the Iranian Chemical Society
Table 3 System suitability data for the proposed TLC-densitometric method for the determination of camyloꢀn-2HCl and paracetamol
Parameter
Camyloꢀn-2HCl
Paracetamol
Ref value [41]
Retardation factor (RF)
0.84
0.19
1
0.72
0.39
1
Capacity factor (K
Tailing factor
̀)
The higher the value, the longer the retention factor
=1 for typical symmetric peak
Number of theoretical plates (N)
Height HETP(H)
864.36
0.008
501.76
0.014
Increase with eꢃciency of separation
The smaller the value, the higher the eꢃciency
in diꢂerent ratios, and it was valid for simultaneous determi-
nation of both drugs with mean recoveries of 100.38%± 1.29
and 99.97% ± 1.59 for camyloꢀn-2HCl and paracetamol,
respectively (Table 5). Densitometric method was stability
indicating for camyloꢀn-2HCl so its selectivity was assessed
by preparing mixtures containing diꢂerent ratios of the
camyloꢀn-2HCl, its degradate and paracetamol with mean
recoveries of 100.85± 1.44 and 99.82± 1.68 for both drugs,
respectively (Table 5).
(d) Application to pharmaceutical formulation
The proposed methods were successfully applied for
the determination of camyloꢀn-2HCl and paracetamol in
®
Anafortan tablet indicating no interference by excipients or
additives (Table 6). The results obtained by the two proposed
methods were statistically compared with those obtained by
a reported method [38] which involves HPLC determina-
tion of both drugs. No signiꢀcant diꢂerence was observed
Table 4 Regression and assay validation parameters by the proposed methods
Camyloꢀn-2HCl
Paracetamol
HPLC
TLC*
HPLC
TLC
λ max (nm)
Linearity range
Slope
Intercept
Correlation coeꢃcient (r )
220
254
220
254
3.0–90.0 (µg/mL)
27.49
15.63
5.0–40.0 (µg/band)
98.48
222.45
10.0–95.0 (µg/mL)
74.97
706.8
0.1–0.5 (µg/band)
11,248
650.8
2
0.9999
0.9993
0.9996
0.9994
*
*
Accuracy (mean%± SD)
Precision (RSD%)
100.67± 1.04
100.59± 0.92
98.47± 0.60
99.08± 2.05
Intraday
Interday
0.66
1.46
0.85
1.16
0.67
1.47
0.65
1.34
Standard addition
100.03± 1.98
100.28± 1.69
98.69± 1.25
100.44± 0.72
*
Average of nine determinations
Table 5 Determination of camyloꢀn-2HCl and paracetamol in their laboratory-prepared mixtures using the proposed methods
HPLC
TLC
camyloꢀn/par-
R % of camyloꢀn
R % of paracetamol
(A:B:C) (µg/band)
% of acidic
degradate
R % of camyloꢀn
R % of paracetamol
acetamol (µg/mL)
5
5
9
3
7
–
–
0:50
:95
0:10
0:70
0:30
100.48
98.92
99.23
101.46
101.82
–
98.22
100.26
98.46
101.84
101.06
–
35:5:0.1
12.5
25
37.5
50
62.5
75
87.5
101.63
99.9
98.05
101.53
101.73
102.23
100.88
100.85± 1.44
99.57
98.69
100.81
101.96
101.64
98.32
30:10:0.3
25:15:0.4
20:20:0.1
15:25:0.3
10:30:0.5
5:35:0.5
–
–
97.72
99.82± 1.68
Mean%± SD
100.38± 1.29
99.97± 1.59
Mean %± SD
R % is the recovery %
A:B:C) is camyloꢀn/acidic degradates/paracetamol
(
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Journal of the Iranian Chemical Society
Table 6 Determination
Camyloꢀn-2HCl
HPLC TLC
Mean%± SD 101.42± 1.32 100.46± 1.55 99.55± 1.28 100.10± 0.86 100.53± 1.63 99.52± 1.13
Paracetamol
of camyloꢀn-2HCl and
paracetamol in pharmaceutical
formulation by the proposed
methods and in comparison
with the reported method [38]
Reported [38] HPLC
TLC
Reported [38]
Variance
Number
t test
1.73
5
2.28
1.05
2.41
5
1.02
1.46
1.65
0.74
5
0.91
0.58
2.67
5
1.13
2.08
1.28
5
–
–
5
–
–
f-test
The theoretical t- and F-values at P=0.05 were 2.31 and 6.3, respectively
Reported [38]—HPLC method for simultaneous determination of camyloꢀn and paracetamol solution
using water C18 column with UV detection at 220 nm and a mobile phase consisting of 0.05% triꢁuoro-
acetic acid in water and 0.05% triꢁuoroacetic acid in acetonitrile (50:50 v/v), at a ꢁow rate of 1.0 mL/min
between the two methods as indicated by t test and f ratio
results (Table 6).
8. N.N. Kadam, P.C. Patil, R.R. Singh, Int. J. Pharm. Pharm. Sci.
3
, 153–158 (2011)
9
.
R.R. Sing, V.M. Rathnam, J. Singh, K.R. Vegesna, ISRN Anal.
Chem. (2012). https://doi.org/10.5402/2012/586415
1
0. S. Rathinam, K.S. Lakshmi, Int. J. Pharm. Pharm. Sci. 6, 585–
5
89 (2014)
Conclusion
11. R.R. Singh, M.V. Rathnam, Am. J. Pharm. Technol. Res. 2,
4
48–457 (2012)
1
1
1
2. P.S. Barde, A.Y. Desai, S.M. Roy, V.V. Vaidya, ACAI J. 10,
The ꢀrst development of stability-indicating TLC densi-
tometric method for the determination of paracetamol and
camyloꢀn-2HCl in the presence of camyloꢀn acidic deg-
radate is produced in this work with full identiꢀcation of
2
07–210 (2011)
3. R.R. Singh, M.V. Rathnam, Anal. Chem. Indian J. 2, 321–331
(2012)
4. R.R. Singh, M.V. Rathnam, S.J. Singh, R.K. Vegesna, Am.
J. Anal. Chem. 2, 944–952 (2011). https://doi.org/10.4236/
ajac.2011.28110
1
camyloꢀn acid degradate by IR and HNMR and a sug-
gestion of degradation pathway which is considered as an
advantage over all the published stability methods. Also
this work introduces a simple, sensitive and accurate HPLC
method for the determination of camyloꢀn and paraceta-
mol mixture with high sensitivity over all published HPLC
methods [37, 38].
1
5. A.M. Saeed, Int. J. Pharm. Sci. Rev. Res. 42, 53–57 (2017)
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Compliance with ethical standards
4
2
21–437 (2017). https://doi.org/10.1080/22297928.2017.13539
1
Conflicts of interest The authors declare no conꢁict of interest.
2
2
2
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