SIMULTANEOUS DETERMINATION OF LEVOCETIRIZINE AND AMBROXOL BY NEW
ANALYTICAL METHODS.
MASTER OF PHARMACY DISSERTATION PROTOCOL,
SUBMITTED TO THE
RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES KARNATAKA,
BANGALORE.
BY
FADADU TRUSHANT DIPAKBHAI
Under The Guidance Of
MRS. PADMAVATHI P. PRABHU
DEPARTMENT OF QUALITY ASSURANCE.
SRINIVAS COLLEGE OF PHARMACY, MANGALORE – 574143.
2010 – 2012
RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES,
BANGALORE, KARNATAKA.
ANNEXURE-II
.PROFORMA FOR REGISTRATION OF SUBJECTS FOR DISSERTATION
1.
NAME OF THE CANDIDATE
AND ADDRESS (IN BLOCK LETTERS)
FADADU TRUSHANT DIPAKBHAI
SRINIVAS COLLEGE OF PHARMACY,
VALACHIL,
POST-FARENGIPETE
MANGALORE-574143,
KARNATAKA.
2.
NAME OF THE INSTITUTION
SRINIVAS COLLEGE OF PHARMACY,
VALACHIL,
POST- FARANGIPETE
MANGALORE-574143,
KARNATAKA
3.
COURSE OF STUDY AND SUBJECT
MASTER OF PHARMACY
(QUALITY ASSURANCE)
4.
DATE OF ADMISSION OF COURSE
31st may 2010.
5.
TITLE OF TOPIC
SIMULTANEOUS DETERMINATION OF LEVOCETIRIZINE AND AMBROXOL BY NEW
ANALYTICAL METHODS.
6.0
7.0
8.0
BRIEF RESUME OF THE INTENDED WORK:
6.1NEED FOR THE SYUDY:
Several spectrophtometric methods have been used for
quantitative determination of levocetirizine and ambroxol in
pharmaceutical formulations and in human plasma.
Levocetirizine and ambroxol is novel in market and no much data
is available on Analytical methods by UV and HPLC method. Our
intension is to develop economic, less time consuming method for
the estimation of levocetirizine and ambroxol by different
analytical methods such as UV and HPLC.
Levocetirizine (as levocetirizine dihydrochloride) is a
third-generation non-sedative antihistamine, developed from the
second-generation antihistamine cetirizine. Chemically,
levocetirizine is the active enantiomer of cetirizine. It is the
L-enantiomer of the cetirizine racemate. Levocetirizine works by
blocking histamine receptors. It does not prevent the actual
release of histamine from mast cells, but prevents it binding to
its receptors. This in turn prevents the release of other allergy
chemicals and increased blood supply to the area, and provides
relief from the typical symptoms of hay fever.
Levocetirizine is called a non-sedating antihistamine as it does
not enter the brain in significant amounts, and is therefore
unlikely to cause drowsiness. However, some people may experience
some slight sleepiness, headache, mouth dryness, vision
problems.
6.2 REVIEW OF LITERATURE:
6.2.1 LEVOCETIRIZINE
Molecular Structure:
Name:
Levocetirizine
Systematic (IUPAC) name:
2-[2-[4-[(R)-(4-chlorophenyl)-phenyl-methyl]piperazin-1-yl]ethoxy]acetic
acid
Molecular Formula:
C21H25Cl
HYPERLINK "http://en.wikipedia.org/wiki/Nitrogen" \o
"Nitrogen"N2O3
Molecular Weight: 388.888
Bioavailability: high
Protein binding:90%
Half life:6 to10 hours
· Shaikh KA and Patil AT1, has given Stability indicating
reverse phase high performance liquid chromatographic method for
the simultaneous determination of Levocetirizine dihydrochloride
and Pseudoephedrine sulfate in pharmaceutical formulation. The
mobile phase A consisted of Potassium dihydrogen phosphate Buffer
0.05M and 1-Ocatne sulphonic acid sodium salt 0.25%, pH adjusted to
3.0 with orthophospheric acid. Mobile Phase B: Acetonitrile,
Gradient elution at flow rate of 1 ml/min and Column temperature at
40oC. Detector wavelength of 242 nm using a photodiode array
detector. The described method shows excellent linearity over a
range of 200–100 μg/ml for Levocetirizine dihydrochloride and
7200-360 μg/ml.1 for Pseudoephedrine sulfate. The correlation
coefficient for Levocetirizine dihydrochloride and Pseudoephedrine
sulfate are 0.9999.
· Ashokkumar S, Senthil RM, Perumal P.2 In this study, reverse
phase high performance liquid chromatographic method have been
developed and validated for the simultaneous determination of
Montelukast and Levocetirizine combined pharmaceutical formulation.
HPLC separation was achieved with a Phenomenex Luna, C18 (250 x 4.6
mm i.d.,5µ) as stationary phase and phosphate buffer (pH adjusted
to 5.5 with dil. KOH): Acetonitrile (65:35, v/v) as eluent, at a
flow rate of 1.5ml/min. UV detection was performed at 230 nm. The
retention time of Montelukast and Levocetirizine were found to 3.7
and 6.1 min respectively. Results of the analysis were validated
statistically by recovery studies.
· Prashant S, Virag S, Dhananjay G, Rohit S, Shital kumar P,
Dhanya Kumar C.,3 given selective, simple, accurate and
reproducible spectrophotometric method for the estimation of
levocetirizine dihydrochloride in bulk and pharmaceutical
formulation. The drug obeyed the Beer’s law and showed good
correlation. It showed absorption maxima at 231nm; in Phosphate
Buffer Solution pH 7.0. The developed method was validated with
respect to linearity, accuracy and precision.
· Lakshmana SP, Shirwalkar AA, Anie S, Dinesh Kumar C, and
Arvind Kumar 4 given a spectrophotometric method that is developed
for simultaneous estimation of ambroxol hydrochloride and
levocetirizine dihydrochloride. The method involved solving
simultaneous equations based on measurement of absorbance at two
wavelengths 242 nm and 231 nm, the λ max of ambroxol hydrochloride
and levocetirizine dihydrochloride, respectively. Beer's law was
obeyed in the concentration range 10–50 μg/ml and 8–24 μg/ml for
Ambroxol hydrochloride and Levocetirizine dihydrochloride
respectively. Results of the method were validated statistically
and by recovery studies.
AMBROXOL
Ambroxol is a secretolytic agent used in the treatment of
respiratory diseases associated with viscid or excessive mucus.
Ambroxol is available in many formulations such as syrup,dry
powder,drops,ampoules etc.
6.2.2 REVIEW OF LITERATURE:
Molecular structure:
Name:
Ambroxol
IUPAC Name:
trans-4-(2-Amino-3,5-dibrombenzylamino)-cyclohexanol
Molecular Formula:
C13H18Br2N2O
Molecular Weight:
378.10
· Neela MB, Santosh KG, and Harinath NM5, gave
spectrophotometric method for the simultaneous analysis of ambroxol
hydrochloride and cetirizine hydrochloride in their tablet
formulation. The chromatographic methods were standardized using a
HIQ SIL-C 18 column (250x4.6 mm i.d., 10 µm particle size) with UV
detection at 229 nm and mobile phase consisting of
methanol-acetonitrile-water (40:40:20, v/v/v). Ambroxol
hydrochloride and cetirizine hydrochloride have absorbance maxima
at 243 nm and 229 nm, respectively. The isoabsorptive wavelength
for both the drugs was 236 nm. For absorbance ratio method
developed, wavelengths selected were 243 nm and 236 nm.
· Heinanen M, Barbas C6 gives the method for estimation of
compounds by HPLC with UV detection at 247 nm in syrup as
pharmaceutical presentation. Optimal conditions were: Column
Symmetry Shield RPC8, 5 micron 250 x 4.6 mm, and
methano1/(H(3)PO(4) 8.5 mM/triethylamine pH=2.8) 40:60 v/v.
Validation was performed using standards and the pharmaceutical
preparation which contains the compounds described above. Results
from both standards and samples show suitable validation
parameters. The pharmaceutical grade substances were tested by
factors that could influence the chemicals.
· Krupa MK, Balasundaram J, Amit P. Khandhar, Rajnish KM 7,
developed and validated analytical method for quantitative
determination of Levofloxacin and Ambroxol hydrochloride in a new
tablet formulation. Chromatographic separations of the two drugs
were analyzed on a Hypersil BDS C18 column (25cm X 4.6mm, 5µm). The
mobile phase constituted of Buffer: Acetonitirile: Methanol
(650:250:100) with triethylamine and pH adjusted to 5.2 with dilute
orthophosphoric acid was delivered at the flow rate 1.0 mL/min.
Detection was performed at 220 nm. Separation was completed within
10 min. Calibration curves were linear with coefficient correlation
between 0.99 to 1.0 over a concentration range of 7 to 22 µg/mL of
Levofloxacin and 50 to 150µg/mL for Ambroxol hydrochloride
respectively. The relative standard deviation (R.S.D) was found
<2.0%.
6.3 OBJECTIVES OF THE STUDY:
· To determine λ max in different solvent and pH condition for
levocetirizine and ambroxol.
· To develop simple, selective, rapid, precise and economical
analytical method for the estimation of levocetirizine and
ambroxol.
· To validate method in terms of accuracy, precision, linearity,
limit of detection, limit of quantitation etc.
MATERIALS AND METHODS:
The standard drug will be requested from the industries in
India. Depending on the availability of facilities, one of the
following methods will be applied for the estimation of
levocetirizine and ambroxol.
1. UV Method:
Standard stock solution can be prepared by dissolving 100mg of
levocetirizine and 100mg of ambroxol in 100 ml of water separately
and then final volume of both solutions made up to 100 µg/ml of
levocetirizine and ambroxol in volumetric flask. Solutions
containing 20 µg/ml of both Levocetirizine and Ambroxol can be
prepared and scanned in UV region separately for λ max.
For the compound levocetirizine and ambroxol λmax can be
obtained, By dilution of Standard drug in 5,10,15,20 ….50 µg/ml of
each drug can be prepared and scanned at 200-400 nm. The values of
absorbances can be recorded. From this data absorptivity and molar
absorbance can be determined for levocetirizine and ambroxol.
Finally, drug formulations can be estimated from the available
data.
2. HPLC Method:
Mixed solutions consisting of 100 µg/ml of levocetirizine and
ambroxol can be prepared in different combinations of solvents
using acetonitrile, water, acetone, methanol. Then, it is passed in
HPLC column using different mobile phases. From HPLC data Retention
time, Peak area can be obtained which is useful for estimation of
drug in formulations.
All the validation parameters such as LOD, LOG, Linearity,
Precision and Accuracy can be validated using ICH Q2b
guidelines.
The above method likely to change depending on the observations
recorded time to time.
7.1 Material:
HPLC instrument: Jasco International Co Ltd., Japan. With C18
column. Detector: UV-visible. Bulk Drugs: Levocetirizine and
Ambroxol,
formulation: combination of Levocetrizine and Ambroxol tablets
capsules syrup, drops etc. Solvents: HPLC grade water, hexane,
ethyl acetate, dichloromethane, methanol, acetonitrile, ethanol,
chloroform, glacial acetic acid, acetone, propanol Chemicals for pH
adjustment: phosphate buffer, ortho-phosphoric acid, hydrochloric
acid, sodium hydroxide, potassium bromide etc.
7.2 Source of data:
Data will be obtained from Pubmed, Science direct, Medline, and
other internet facilities, literature search and related articles
from library of Srinivas College of Pharmacy, Mangalore.etc.
7.3 Method of Collection of Data (Including Sampling Procedure,
If Any)
1. Procurement of the drug and marketed formulations.
2. Solubility determination of Levocetirizine and Ambroxol in
different solvents and buffers.
3. Determination of wavelength of maximum absorbance and
intensity of absorption in different solvents and buffers for
various analytical studies.
4. Develop and validate analytical method for Levocetirizine and
Ambroxol by UV-Visible spectrophotometer using standard
absorptivity value, standard calibration curve, single point and
double point standardisation. Develop and validate HPLC method to
estimate Levocetirizin and Ambroxol in bulk and pharmaceutical
formulations.
7.4 Does the study require any investigation or intervention to
be conducted on patients or other humans or animals? If so, please
mention briefly.
--NOT APPLICABLE--
7.5 Has ethical clearance been obtained from your institution in
case of 7.3?
--NOT APPLICABLE--
LIST OF ReferenceS:
1. Shaika KA, Patil AT. Stability indication LC methods for the
simultaneous determination of levocetirizine and pseudoephedrine
sulphate in tablet dosage forms. Int J ChemTech Res 2010;
2(1):454-61.
2. Ashokkumar S, Senthil RM, Perumal P. RP-HPLC method
development and validation for simultaneous estimation of
montelukast sodium and levocetirizine dihydrochloride. Int J Pharma
Res 2009; 1(4):8-12.
3. Prashant S, Virag S, Dhananjay G, Rohit S, Shital kumar P,
Dhanya Kumar C. Validation of UV Spectrophotometric method for
estimation of levocetrizine HCl in Bulk and Pharmaceutical
Formulation. Ind J Pharma Res 2010; 3(10):2386-87.
4. Lakshmana SP, Shirwalkar AA, Anie S, Dinesh Kumar C, Arvind
Kumar. Simultaneous UV Spectrophotometric estimation of ambroxol
HCl and Levocetrizine HCl. Ind J Pharma Sci 2008; 70(2):236-38.
5. Neela MB, Santosh KG, Harinath NM. Spectrophotometric
estimation of ambroxol HCl and cetrizine HCl in tablets. Asian J
Pharm 2008; 2(3):159-62.
6. Heinanen M, Barbas C. Validation of an HPLC method for the
quantification of ambroxol hydrochloride and benzoic acid in syrup
as pharmaceutical form stress test for stability evaluation. J
Pharm Biomed Ana 2001; 24:1005–10.
7. Krupa MK, Balasundaram J, Amit P. Khandhar, Rajnish KM.
Quantitative determination of levofloxacin and ambroxol HCl in
pharmaceutical dosage form by RP-HPLC. Eura J Anal Chem 2007;
2(1):21-30.
9.
Signature of the candidate
(FADADU TRUSHANT )
10.
Remarks of the Guide
The candidate is working under my direct supervision in
laboratories of Srinivas College of Pharmacy, Mangalore-574143.
11.
11.1 Name and Designation of Guide
11.2 Signature
MRS. PADMAVATHI P. PRABHU M.Pharma (Ph.D)
ASSISTANT PROFESSOR
(DEPARTMENT OF QUALITY ASSURANCE)
SRINIVAS COLLEGE OF PHARMACY
MRS. PADMAVATHI P. PRABHU
11.3 Head of the department
11.4 Signature
PROF. DR. E.V.S SUBRAMANYAM M.Pharma Ph.D
DEPARTMENT OF QUALITY ASSURANCE,
SRINIVAS COLLEGE OF PHARMACY.
DR. E.V.S. SUBRAMANYAM
12.
12.1 Remarks of Principal
12.2 Signature
FORWARDED AND RECOMMENDED FOR FAVOURABLE CONSIDERATION.
PROF. A.R. SHABRAYA
