DEVELOPMENT AND VALIDATION OF NOVEL ANALYTICAL METHOD FOR SIMULTANEOUS ESTIMATION OF PARACETAMOL AND

PAMABROM IN BULK AND PHARMACEUTICAL DOSAGE FORM

M. Pharm Dissertation Protocol Submitted to

Rajiv Gandhi University of Health Sciences, KarnatakaBangalore – 560 041

ByMr. DEBA PRAKASH BORUAH B. Pharm.

Under the Guidance ofMr. SYED NIZAMUDDIN M. Pharm.

Asst. ProfessorDepartment of Quality Assurance

Acharya & B.M. Reddy College of Pharmacy,Soldevanahalli, Bangalore - 560 090,

2011– 2013.

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RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES, KARNATAKA

BANGALORE.

ANNEXURE – II

PROFORMA FOR REGISTRATION OF SUBJECTS FOR M.PHARM DISSERTATION

1.0 Name of the candidate and

address DEBA PRAKASH BORUAH

S/O MR. AMRIT CH. BORUAH

TINGALI SEN SWA GAON P.O: CHIRAKHUNDA

DIST: SIVASAGAR ASSAM, PIN: 785666

2.0 Name of The institution ACHARYA & B.M. REDDY COLLEGE OF

PHARMACY,

Soldevanahalli, Bangalore-560 090.

3.0 Course of study and subject M. PHARMACY (QUALITY ASSURANCE)

4.0 Date of admission 12 December,2011

5.0 Title of the topic

DEVELOPMENT AND VALIDATION OF NOVEL ANALYTICAL METHOD FOR

SIMULTANEOUS ESTIMATION OF PARACETAMOL AND PAMABROM IN BULK

AND PHARMACEUTICAL DOSAGE FORM.

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6.0 BRIEF RESUME OF THE INTENDED WORK:

6.1:- NEED FOR PRESENT STUDY :

The introduction of large number of newer drugs and their formulation may also lead to

widespread distribution of substandard or even counterfeit drugs and their formulations

in the market. Quality control and quality assurance of pharmaceutical chemicals and

their formulation are essential for ensuring the general public against the hazards of

substandard drugs. Pharmaceutical analysis is indispensable in the process of quality

control for statutory certification of drugs and their formulation either by the industry or

by the regulatory authorities. Thus constant development of new and improved analytical

methods for accurate determination of drugs in raw materials and in pharmaceutical

dosage forms is essential for quality control studies.

One such drug combination of Paracetamol and Pamabrom in tablet dosage form is used

for analgesic and diuretic action1. On literature survey, it was found that very few

analytical methods reported for the estimation of Paracetamol and Pamabrom in

pharmaceutical tablet dosage form. The present study is to make an attempt to establish

sensitive and accurate methods for the estimation of titled drugs in bulk and tablet

dosage form.

The proposed methods have to be validated for linearity, accuracy, precision, robustness,

limit of quantitation as per ICH guidelines2 for the intendant analytical application.

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DRUG PROFILE

PARACETAMOL 3,4:

Paracetamol is an anti-pyretic and analgesic compound.

Mol. Formula : C8H9NO2

Mol. Wt. : 151.17 g/mol

Solubility : Soluble in water and alcohol

Description : White crystalline powder

pKa : 9.5

PAMABROM 5,6:

It is a substance being used for diuretic activity.

Mol.formula : C7H7BrN4O2

Mol. Wt. : 348.20 g/mol

Solubility : Soluble in water

Description : White to off white amorphous powder

pKa : 3.15

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6.2 Review of literature:

The simultaneous UV spectrophotometric method for the estimation of Paracetamol and

Nimesulide in tablet dosage form was carried out by Kirtawade R et al7. The method

involved was the formation of simultaneous equation of the analytical wavelengths for

the respective drugs in methanol and distilled water (50:50 v/v).

The two constituents of the compounds namely Acetaminophen and Pamabrom tablets in

human plasma by HPLC were determined by Zhou L et al8. The chromatograph was

carried out on Diamon-sil™ C18 using Theophylline as internal standard. The mobile

phase was 0.2% trifluoroacetic acid - acetonitrile (87:13 v/v).

One of the valid analytical method was developed by B Elena and coauthors for the

assay and identification of Pamabrom, Pyrilamine maleate and Ibuprofen. They used

mixture of methanol: acetonitrile: acetic acid of 1.5% in water (30:30:40 v/v) 9 .

A simultaneous estimation of Aceclofenac, Paracetamol and Chloroxazone in tablets was

carried out by G Garg and coauthors. The estimation method was based on the additivity

of absorbances for the determination of Aceclofenac, Paracetamol and Chloroxazone.

The wavelengths for the drugs were found to be 276 nm, 248 nm and 282 nm

respectively in the concentration range of 2-20 µg/mL10.

Establishment of Aceclofenac and Paracetamol in tablet dosage form using reverse phase

HPLC method was determined by Godse VP et al11. The mobile phase used was a

mixture of methanol and water (70:30 v/v).

Gopinath R et al12, estimated Paracetamol and Aceclofenac tablets using RP HPLC

method. The method was carried out on a Hichrom C18 column with a mobile phase

consisting of acetonitrile: 20mM phosphate buffer (pH 5.0) (60:40 v/v).

Li J et al13, investigated the pharmacokinetics of Pamabrom and Acetaminophen tablets

in healthy female volunteers. The blood samples were collected and the serum

concentration was determined by a developed and validated HPLC method.

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6.3 OBJECTIVES OF THE STUDY:

Literature survey reveals that only few analytical methods had been developed for the

simultaneous estimation of Paracetamol and Pamabrom in pharmaceutical dosage. In view of the

above fact the following attempts shall be made.

To develop and validate simple, precise and accurate methods for the simultaneous

estimation of Paracetamol and Pamabrom in bulk dosage form and Pharmaceutical tablet

dosage form by using HPLC using various solvents and buffers.

To validate the method developed by using various analytical parameters such as

accuracy, precision, specificity, robustness, ruggedness, linearity and range as per ICH

guidelines.

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7.0 MATERIALS AND METHODS:

7.1:-SOURCE OF DATA:

The data will be obtained from:

1. JOURNALS :

Indian Journal of Pharmaceutical Sciences.

International Journal of Pharmaceutical Sciences.

International Journal of Drug Development Research.

International Journal of Chemical And Analytical Sciences.

Indian Journal of Chemical Technology.

Journal of Pharmaceutical and Biomedical Analysis.

The Pharmaceutical and Health Science Journal.

2. Helinet of RGUHS.

3. www.google.com

4. www.wikepedia.com

5. www.pubmed.com

7.2:- METHOD OF COLLECTION OF DATA:

Bulk drug samples of simultaneous estimation of Paracetamol and Pamabrom shall be

procured from industries involved in manufacture of them or from suppliers.

The analytical method development, validation study will be carried out in the

Department of Quality Assurance, Acharya & B.M Reddy College of Pharmacy,

Bangalore-90.

Methodology of the study:

HPLC method for simultaneous estimation of Paracetamol and Pamabrom in Bulk

powder & Tablet dosage form will be developed, the following factors will be decided:

Stationary phase, mobile phase, gradient or isocratic elution, wavelength of light for

monitoring the effluent etc.

Novel analytical method for simultaneous estimation of Paracetamol and Pamabrom

will be developed by any one of the following Spectrophotometric methods:

a) First order derivative method.

b) Simultaneous equation method.

c) Q-analysis method (absorption ratio method).

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LC and UV/Vis spectrophotometric method will be Validated, as per ICH Guidelines,

for following parameters:

1) PRECISION:

The response to repeated injections of the same standard solution should have RSD

≤ 1.0%.

Intra assay: Injection of sample from each solution should have RSD ≤ 2.0%.

Intermediate precision: Assay the same sample several times within two to three

days using same experimental conditions but a different analyst and instrument,

RSD ≤ 2.0%

2) ACCURACY:

Direct comparison to reference standard.

Performing recovery studies by spiking the blank matrix of the sample at different

level (50, 75,100,125,150%) of the known level in the sample. Average recovery of

the analyte should be in the range of 98.0-102.0%.

3) ROBUSTNESS:

Changing conditions slightly such as temperature ±5ºC, buffer pH±0.5, and ionic

strength of buffers and level of additives to mobile phase.

The method must be robust enough to withstand such slight changes and allow

routine analysis of the sample.

4) RUGGEDNESS:

Measuring the concentration of the analyte using different parameters.

Different laboratories and different operators and equipments in the same lab.

Different source of reagent and solutions and column.

5) LINEARITY:

Linearity is the degree to which the response measured by the method is directly

proportional to the concentration of the analyte.

By covering the range 0-150% of the expected level of the analyte.

The method must exhibit linearity in the described range.

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6) RANGE :

Verifying that the analytical method provides acceptable precision, accuracy.

Linearity when applied to samples containing analyte at the extremes of the range

as well as within the range

7) SPECIFICITY:

Injecting the sample such as Paracetamol and Pamabrom with Rs not less than 3.0.

Injecting separately different components of the matrix of the sample (tablets)

should clearly separate from analyte with Rs ≥ 2.0.

Tablets will be analyzed by the HPLC and UV/visible spectrophotometric method.

7.3:- DOES THE STUDY REQUIRE ANY INVESTIGATION OR INTERVENTIONS

TO BE CONDUCTED ON THE PATIENTS OR OTHER HUMAN/ANIMALS? IF

SO, PLEASE DESCRIBE BRIEFLY.

- Not Applicable -

7.4:- HAS ETHICAL CLEARANCE BEEN OBTAINED FROM YOUR INSTITUTION

IN CASE OF 7.3?

- Not Applicable-

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8.0 LIST OF REFERENCES:

1. Paracetamol and Pamabrom drug profile [ Cited 2012 Apr 26 ].Available

from:http://indianhealthservices.in/new_approval.php

2. ICH topic Q 2 B Validation of analytical procedures-methodology [internet].1996[cited

2012 May 7].Available from:http://www.Uam.Es/Personal_Pas/Txrf/Mu5.Pdf.

3. Indian Pharmacopeia .Delhi, Govt of India. Ministry of Health & Family Welfare 1996;

2:P554-55.

4. Paracetamol drug profile [Cited 2012 Apr.26] Available from:

http://en.wikipedia.org/wiki/Paracetamol.

5. US Pharmacopoeia 34 NF 29. The United States Pharmacopoeial Convention, Rockvill,

2011; 3:3800.

6. Pamabrom drug profile [Cited 2012 May 8] Available from:

http://en.wikipedia.org/wiki/Pamabrom.

7. Kirtawade R,Salve P, Seervi C, Kulkarni A, Dhabale P. Simultaneous UV Spectroscopic

method for the estimation of Paracetamol and Nimesulide in tablet dosage form. Int J

Chem Tech Res 2010 Apr-Jun; 2(2):818-21.

8. Zhou L, Gu L, Wang Y, Linang J. HPLC for the determination of two constituents in

compound Acetaminophen and Pamabrom tablets in human plasma.Chin J New Drug

Clin Remed 2007.(DOI:-CNKI:SUN:XYYL.0.2007-03-008 )

9. Rodriguez E, Rosas M , B Elena .An unique LC for the assay and identification of

Pamabrom,Pyrilamine maleate and Ibuprofen in softgels formulated in a hydrophilic

solution.[Cited on 2012 May 8].Available from

http://www.aapsj.org/abstracts/Am_2005/AAPS2005_002940.pdf

10. Garg G, Saraf S. Simultaneous estimation of Aceclofenac, Paracetamol and

Chlorzoxazone in tablets. Ind J Pharm Sci. 2007; 69(5): 692-94

11. Godse VP, Deodhar MN, Bhosale AV, Sanowane RA, Sakpal RS, Borar DD et al.

Reverse phase HPLC method for the determination of Aceclofenac and Paracetamol in

tablet dosage form. Asian J Res Chem. 2009 Jan – Mar; 2(1):37-40.

12. Gopinath R, Rajan S, Meyanathan SN, Krishnaveni N, Suresh BA. RP-HPLC method for

the simultaneous estimation of Paracetamol and Aceclofenac in Tablets. Ind J Pharm Sci.

2004; 66(5):694-6.

13. Li J, Ma T, Huo Q, Ziang J, Dong H, Gao S et al . Investigation on the pharmacokinetics

of compound Acetaminophen and Pamabeom tablets in healthy female volunteers. Chin

J Hosp Pharm.2010 .(DOI:-CNKI:SUN:ZGYZ.0.2010.-04-009 )

9.0 SIGNATURE OF THE 11

CANDIDATE

10.0 REMARKS OF THE GUIDE Recommended for Research

11.0 NAME AND DESIGNATION OF:

11.1 GUIDE Mr. Syed Nizamuddin Assistant professor. Department of Quality Assurance,Acharya & B.M. Reddy College of Pharmacy, Soldevanahalli, Bangalore-90

11.2 SIGNATURE

11.3 CO-GUIDE NIL

11.4 SIGNATURE NIL

11.5 HEAD OF THE DEPARTMENT

Mr. A. Cendil Kumar, Professor & HeadDepartment of Quality Assurance,Acharya & B.M. Reddy College of Pharmacy, Soldevanahalli, Bangalore-90

11.6 SIGNATURE

12.0 12.1 REMARKS OF THE PRINCIPAL

12.2 NAME OF THE PRINCIPAL

Dr. Divakar GoliPrincipal,Acharya & B.M. Reddy College of Pharmacy, Soldevanahalli,Bangalore-90.

12.3 SIGNATURE

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