METHOD DEVELOPMENT AND VALIDATION FOR SIMULTANEOUS ASSESSMENT OF NAPROXEN AND PARACETAMOL IN MIXED TABLET DOSAGE FORM BY RP-UPLC
K.KANAKAPARVATHI*, Vijay Nagarjan, Santha Arcot and CH Hemanth Kumar.
 
ABSTRACT
An advancement design and corroboration for simultaneous assessment of Naproxen (NAP) and Paracetamol (PAR) in merged tablet dosage form by UPLC. The column used in determination was C18 thermo fisher (50cm x 4.6 mm x 3µm), mobile phase used in this method was 0.4% ammonium acetate buffer: methanol: acetonitrile (40:40:20), the retention time was about 1.9 minutes and 3 minutes for PAR and NAP of a total run time of 5 minutes, with flow rate of 0.2ml per minute respectively at a wavelength of 271nm, linearity of the method was linear over the range of 38.496 to 57.664μg/ml for Paracetamol and 64.096 to 95.968μg/ml of Naproxen respectively with a correlation of 0.999 for simultaneous assessment for PAR and NAP thus the method was fast, simple, elegant and less time consuming method
Keywords: Naproxen, Paracetamol RP-UPLC, Method validation
INTRODUCTION
Naproxen is chemically 2-Naphthaleneacetic acid, 6-methoxy-α-methyl-(s)-(+)-(s)-6 methoxy-α-methyl-2-naphthaleneacetic acid as shown in (Figure 1). It is a non-steroidal anti-inflammatory drug commonly used for minimizing of moderate to severe torment, delirium, inflammation and stiffness. [6-11].
Paracetamol (PAR) is chemically N-(4-hydroxyphenyl) acetamide (Figure 2), It has analgesic and antipyretic activity for the therapy of subsidiary, non-inflammatory conditions of patient who were prone to gastric symptoms [12-14].
The merger of these two drugs are used in the remedy [11] of Musculoskeletal Disorder (Sprain/Strains) Trauma Fractures/injuries), Occupational affliction, Joint torment, Low Back laceration the literature review supports legion UPLC methods for the evaluation of NAP and PAR independently and in combination with other drugs but There was no UPLC method had been reported for the determination of NAP and PARA in merged dosage form
So an experiment was taken to expand and corroborate a rapid RP-UPLC method [1-5] for the determination of NAP and PARA in mixed tablet dosage forms.

Figure 1 NAPROXEN Figure 2 Paracetamol
 
MATERIALS
NAP and PAR was earned from Ideal analytical and research institution puducherry, India. All chemicals worn were analytical standard. The pharmaceutical tablet dosage form used in this study was NAPROSYN P with a label claim of NAP 300mg and PAR 500mg were purchased from local pharmacy.
INSTRUMENTATION AND APPARATUS
The uplc system used for advancement design and corroboration was thermo accela equipped with 1050 quaternary pump auto sampler and photodiode array detector. The detector output were recorded and processed using chrome quest software version 5.0 sonicator (PCI bath sonicator ) was used for degassing of mobile phase and sonication of the solutions prepared
SOFTWARE:
The statically calculation for the analysis was performed by using Microsoft excel 2010 software (Microsoft, USA)
METHOD CORROBORATION:
SYSTEM SUITABILITY:
System suitability was determined by injecting the standard solution and observed the parameters like retention time, peak area, relative standard deviation, tailing factor, USP theoretical plates.
LINEARITY
For testing of linearity five different concentration of sample solution (80%, 90%, 100%, 110%, and 120%) was injected and checked over by plotting the graph as peak area verses concentration thus the data treated by linear regression analysis.
ACCURACY
Accuracy can be done by injecting the sample solution with known standard concentration and the amount of percentage recovery gives the accuracy of sample.
PRECISION
Precision can be evaluated by Interday and intraday, were the same sample solution has to be assayed for the same day and on different days at different time intervals
ROBUSTNESS
The determination of robustness can be done by changing the experimental condition deliberately. The condition may include of changing in mobile phase flow rate, pH and temperature, the percentage of RSD, tailing factor, resolution, were cross check with the original data.
RESULT DISCUSSION:
The method has validated according to the norms of international harmonization of conference (ICH) guidelines with regards of system suitability, linearity, accuracy, precision and robustness as follows
SYSTEM SUITABILITY
The system suitability tests were carried out to evaluate the resolution and reproducibility of the system for the analysis. The results of the system suitability test were summarized in Table No.1.
Table 1: System suitability results

S.No

PARAMETERS

PAR

NAP

1

Retention Time

1.807

3.007

2

Peak area

410801

306340

3

Percentage area

57.28

42.72

4

Theoretical plates

2633

3306

5

Resolution

0.0000

0.85712

6

Tailing factor

1.754

1.696

Solution stability
The solvents which had been used in the mobile phase were cost effective than the solvents used in the other UPLC methods which are reported in the literatures.
Standard and samples solution stability was studied above 12 and 24 hours and found stable against the freshly prepared standard.
Table2. Results of Solution stability

Time (hrs)

Percentage Assay

Percentage difference in assay

 

PAR NAP

PAR NAP

Initial

99.92

99.99

0.002

0.001

After 12 hrs

99.52

99.57

0.003

0.002

After 24 hrs

99.12

99.19

0.001

0.002

LINEARITY
Linearity of the method was evaluated at 5 different concentration levels of 38.496 to 57.664μg/ml for Paracetamol and 64.096 to 95.968μg/ml of Naproxen respectively. Both the drugs were found to give linear detector response in the concentration under study with correlation coefficient of 0.997 and 0.999 for PAR and NAP respectively.
Table3: Linearity study for NAP and PAR

S.NO

PARAMETERS

PAR

NAP

1

Linearity range

38.49 – 57.664μg/ml

64.09 -95.96μg/ml

2

Correlation coefficient (r2)

0.997

0.999

3

Slope

3769.8726

2867.1591

4

Intercept

1567.7362

0.1591

ACCURACY
Accuracy of the method was determined by recovery test. The percentage recovery was found to be within the concentration of 100 to 115 as 100, 105, 110, and 115 (Table4). All results indicate that the method is highly accurate.
Table: 4(a) accuracy data for PAR

S.NO

ACCURACY LEVEL

STANDARD AREA

SAMPLE AREA

Mg/tab

PERCENTAGE

1

100

404871

393726

499.83

99.97

2

105

404871

413927

525.48

105.1

3

110

404871

433143

549.87

109.97

4

115

404871

454077

576.46

115.29

Table 4(b) accuracy data for NAP

S.NO

ACCURACY LEVEL

STANDARD AREA

SAMPLE AREA

Mg/tab

PERCENTAGE

1

100

306460.4

303506

299.26

99.75

2

105

306460.4

319467

315.00

105.00

3

110

306460.4

334246

329.57

109.86

4

115

306460.4

350847

345.94

115.31

PRECISION
This method was validated for its inter-day and intra-day precision. The results (table4) obtained were within the acceptable limit.
Table 5: results for precision studies

s.no

Parameter(units)

PAR

 

NAP

 

 
 

STANDARD AREA

SAMPLE AREA

PERCENTAGE

STANDARD AREA

SAMPLE AREA

PERCENTAGE

1

Interday precision
(1st day)
(2nd day)
(3rd day)

404871
404871
404871

401886
402568
403442

100.87
99.28
100.74

306460
306460
306460

307076
307209
309589

99.77
98.08
100.07

2

Intraday precision
1sthrs
2nd hrs
3rd hrs

404871
404871
404871

402645
401507
400271

100.17
100.65
100.49

306460
306460
306460

309957
307438
307946

99.82
99.76
100.07

3

Average

 
 

100.366

 
 

99.595

4

SD

 
 

0.584

 
 

0.75

5

RSD

 
 

0.582

 
 

0.758

ROBUSTNESS
The robustness of the method was determined and the percentage RSD of the results was found to be less than 2.0%, which demonstrate that the developed method is robust.
Table6. Results of Robustness parameter

 
 

CHANGED PARAMETERS

 
 

FLOW RATE

WAVE LENGTH

S.NO

PARAMETERS

190

210

269

273

 
 

PAR

NAP

PAR

NAP

PAR

NAP

PAR

NAP

1

Retention time

1.938

3.215

1.70

2.832

1.810

3.005

1.810

3.007

2

Area

462947

347334

406134

306784

432154

322852

426295

347442

3

% area

57.13

42.87

56.97

43.03

57.24

42.76

55.10

44.90

CONCLUSION:
Thus, the above stated method for determination of PAR and NAP by UPLC method concludes as it can be quantified simultaneously by using of isocratic mobile phase of 0.4% ammonium acetate buffer: methanol: acetonitrile (40:40:20), by using of PDA detector at 271 nm. Thus the proposed method is simple, precise, accurate, rapid and sensitive, where it can be applied successfully for the assessment of PAR and NAP in combined pharmaceutical formulations.
ACKNOWLEDGEMENT
The authors are thankful to ideal analytical and research laboratory pondycherry, India for all the facilities provided to complete our work.