Estimation of Metaxalone in Bulk and in Tablet Dosage Form by RP-HPLC

 

D Nagavalli*, ASK Sankar, K Anandakumar, T Vetrichelvan and M Balaji

Dept. of Pharmaceutical Analysis, Adhiparasakthi College of Pharmacy, Melmaruvathur, T.N., India. 603319.

*Corresponding Author E-mail: d_nagavalli@yahoo.co.in

 

ABSTRACT:

A simple, selective, rapid and precise RP-HPLC method for the estimation of Metaxalone  in bulk material and in pharmaceutical formulation has been developed and validated. An isocratic elution at a flow rate of 1.0 ml/min was employed on a Phenomenax Luna C18 column (150 X 4.6 mm i.d., 5 m) at ambient temperature. The mobile phase consisted of Phosphate buffer: pH (5.0) Acetonitrile: Methanol in the ratio of (40:40:20). The UV detection wavelength was documented at 280 nm. The retention time Metaxalone was found to be 4.19 min. The method obeys Beer's Law in the concentration range of 20-100 mg/ml. The method was validated as per standard analytical procedures. The correlation co-efficient value was found to be 0.9997. The limit of detection and the limit of quantification were found to be 0.0.3205 mg/ml and 0.9712 mg/ml respectively. The amount of Metaxalone present in formulation was found to be 399.98 mg. The method was validated statistically and by recovery studies. Hence, the proposed method can be proficiently applied for the routine analysis of Metaxalone in bulk and in tablet formulation.

 

KEYWORDS: Metaxalone, RP-HPLC, Skeletal muscle relaxant, Skelaxin

 


INTRODUCTION:

Metaxalone is an Skeletal Muscle Relaxant drug and chemically it is 5-[(3,5-dimethyl  phenoxy) methyl]-2-oxazolidinon. The reported method1 for Metaxalone was done only in plasma by liquid chromatography coupled tandem mass spectrometry. Tablet dosage form of Metaxalone was introduced recently and yet no method is reported for the estimation of the same. So a simple, selective, rapid, precise and accurate RP-HPLC method for the estimation of Metaxalone in bulk material and pharmaceutical formulation has been developed and validated 2,3.

 

STRUCTURE:

 

MATERIALS AND METHOD:

Drug Sample:

Metaxalone was obtained as a gift sample from AURABINDO Pharmaceuticals Pvt.Ltd., Hyderabad, India. The drug sample received was authenticated from its melting point.

 

Formulation used

Skelaxin tablets containing 400 mg of Metaxalone was purchased from ATLANTA pharmacy U.S.A

 

Reagents and Chemicals:

All the chemicals used were of HPLC grade and procured from Qualigens India Ltd., LOBA and Ranbaxy fine Chemicals Ltd.,India.

 

Instruments Used:

Selection of absorption maximum for the drug Metaxalone in the mobile phase were made on a Shimadzu 1700 double beam UV – Visible spectrophotometer. The   pH of the mobile phase was adjusted by using Elico pH meter Li 610. The Shimadzu electronic balance (AX-200) was used for the weighing purpose.  The liquid chromatographic system used was Shimadzu HPLC model containing LC -1043 pump, variable wavelength programmable UV/VIS detector.  Chromatographic analysis was performed using Winchrom software. C18 Phenomenax Luna

Column was used for isolation.

 

METHOD - RP-HPLC:

Method:

Selection of Chromatographic method:

Proper selection of the method depends upon the nature of the sample, molecular weight, and solubility. The drug selected for the present study was polar, so reversed phase chromatography can be used. C18 column was chosen as stationary phase with different compositions of mobile phase such as acetonitrile, methanol and phosphate buffer were used.

 

Selection of mobile phase and λmax:

A solution of Metaxalone 10 µg/ml was scanned in the UV region using the solvent, Phosphate buffer: Acetonitrile: Methanol in the ratio of 40:40:20. The λmax was found at 280 nm. This wavelength was followed for the estimation of Metaxalone by RP- HPLC method.

 

Chromatographic condition:

Optimized chromatographic parameters were used for the estimation of Metaxalone

Mode of operation        :          Isocratic

Stationary phase           :          C18

Mobile phase                :          Phosphate buffer (pH-5.0)  

                                                 : Acetonitrile: Methanol

Ratio                             :          40 : 40 : 20

Detection wavelength  :          280 nm.

Flow rate                      :          1 ml/min

Load                             :         20 ml.

 

Preparation of standard solution:

An accurately weighed quantity of Metaxalone was dissolved in a minimum quantity of methanol the total volume was brought to 50 ml with methanol (200 µg/ml).

 

Linearity and calibration:

From the standard solution 1-5 ml of 200 µg/ml transferred into 10 ml volumetric flask and made upto the mark to obtain the concentration range from 20-100 µg/ml and the calibration curve was plotted between concentration versus peak area.

 

Quantification of Metaxalone in formulation:

Each tablet containing 400 mg of Metaxalone was taken. The average weight of each tablet were found and powdered. The powdered was weighed equivalent to 250 mg of Metaxalone and was transferred into a 50 ml volumetric flask, added sufficient quantity of methanol and were sonicated for few minutes and made up to the mark with methanol. The solution was filtered through Whatmann filter paper No.41. From these clear solution further dilutions were made with mobile phase and produced 50 µg/ml solution. The peak area measurements were done by injecting the sample (50 µg/ml) six times and the amount of Metaxalone calculated from the respective calibration curve

 

Recovery studies:

To ensure the reliability of the method, recovery studies were carried out by mixing a known quantity of standard drug solution with the pre-analyzed sample formulation and the contents were mixed and made to the volume with mobile phase and re-analyzed by the proposed method. The percentage recovery was calculated4. The proposed method was validated and found that the excipients and additives did not interfering the developed method. The proposed methods were validated statistically6.

 

Limit of detection and Limit of quantification:

Preparation of calibration curve from the serial dilutions of standard was repeated for six times. The limit of detection and limit of quantification was calculated by using the average value of slope and standard deviation response.

 

RESULTS AND DISCUSSION

An effort has been made to identify a simple, cost effective, economic, specific and accurate method for the estimation of Metaxalone in formulations.

 

The mobile phase used which consists of Phosphate Buffer (pH- 5): Acetonitrile: Methanol was chosen in the ratio of 40:40:20 and the lmax of Metaxalone in mobile phase were found at 280 nm and the UV spectrum was shown in fig-1. The flow rate was kept at 1.0 ml/minute. Various concentration of raw material such as 20-100 mg/ml of Metaxalone were prepared, injected and the Calibration Curve were shown fig-2 The optical characteristics such as Beer’s law limits (20-100 mg/ml), correlation coefficient 0.9997, slope 41553.5485 and intercept 32629.2381. Which were shown in Table-1.The Limit of Detection and the Limit of Quantification were determined from the linearity studies which have done six times and then it was calculated by using slope and standard deviation response.  The LOD was found to be 0.3205 mg/ml and the LOQ was found to be 0.971288 mg/ml.   Which were shown in Table-1

 

Fig-1: Ultra violet absorption spectrum of Metaxalone using Mobile phase

Relation between concentration and absorbance

 

Table-1: OPTICAL CHARACTERISTICS OF METAXALONE BY RP-HPLC METHOD

Parameters

Method

lmax(nm)

280nm

Beers law limit (mg/ml)

20-100 µg/ml

Correlation coefficient (r)

0.9997

Régression équation (y=mx+c)

Y=41553.5485+32629.2381

Slope(m)

41553.5485

Intercept(c)

32629.2381

LOD (mg/ml)

0.320525186

LOQ (mg/ml)

0.971288443

Table -2: SYSTEM SUITABILITY PARAMETERS FOR THE OPTIMIZED CHROMATOGRAM BY RP-HPLC METHOD

S.NO.

PARAMETERS

METAXALONE

1.

Tailing factor

1.27

2.

Asymmetrical factor

1.53

3.

Theoretical plates

5222

4.

Theoretical Plate Per unit length

348.13

 

The system suitability test parameters5 such as Theoretical plates (5222), Tailing factor (1.27), Asymmetric factor (1.53) and Capacity factor (1.92) were calculated. The parameters were found to be satisfactory as per guidelines. Which were shown in Table-2

 

Fig-2:Calibration curve for HPLC method using mobile phase

 

Injection repeatability was performed for same concentrations six times to the formulation skelaxin and the amount present was found to be 399.98 mg.  Which were shown in  fig-3 and Table-3.

 

Fig-3: Chromatogram for analysis of Metaxalone  Formulation

 

Table-3: QUANTIFICATION OF FORMULATION SKELAXINBY RP-HPLC METHOD

S. No

Label Claim

(mg)

Amount found (mg)

Percentage obtained

Average

(%)

S.D.

% RSD

1.

2.

3.

4.

5.

6.

400

401.72

100.43

99.99

0.3656

0.3656

400.80

100.20

399.76

99.94

399.20

99.80

397.60

99.40

400.80

100.20

Accuracy5 was confirmed by recovery studies by adding known amount of pure drug to the previously analyzed formulations and the mixture was re-analysed by the proposed method. The % recovery of skelaxin was found to be 100.05 %. The proposed method was validated and found that the excipients and additives did not interfering the developed method. Which were shown in Table-4

 

 

Table -4: RECOVERY STUDIES FOR FORMULATION: SKELAXINBY RP-HPLC METHOD

S.NO.

 

Con.

(%)

Amount  Present µg/ml)

Amount Added

 (µg/ml)

Amount Estimated (µg/ml)

Amount recovered (µg/ml)

% Recovery

 

S.D.

 

% RSD

 

1

50

24.99

25

49.5

24.51

98.04

1.21

1.209

2

50

24.99

25

49.86

24.87

99.48

3

50

24.99

25

50.19

25.2

100.8

4

75

24.99

50

75.23

50.24

100.48

5

75

24.99

50

74.47

49.48

98.96

6

75

24.99

50

75.43

50.44

100.88

7

100

24.99

75

101.17

76.18

101.57

8

100

24.99

75

99.48

74.49

99.32

9

100

24.99

75

100.5

75.51

100.68

 

All the above parameters combined with the simplicity and ease of operation ensures that the application of proposed method in the assay of drug in pharmaceutical dosage forms.  Thus the proposed HPLC method may also be applied for the estimation of Metaxalone raw materials and Pharmaceutical dosage form.

 

ACKNOWLEDGEMENTS:

The authors are thankful to Arulthiru Bangaru Adigalar, President, Thirumathi Lakshmi Bangaru Adigalar, Vice President and Dr.T.Ramesh, Managing Director MAPIMS, Melmaruvathur for providing necessary facilities for carry out this work.

 

REFERENCES:

1.       Nirogi, R.V. Kandikere, V.N. Shukla, M. Mudigonda, K. Shrivastava, W. Datla, P.V. Quantification of Metaxalone in human plasma by Liquid chromatography coupled to tandem mass spectrometry.  Journal of Analytical Toxicology, Florida, (Vol-30), 4, 2006, 245-251.

2.       ICH, Q2A, Text on Validation of Analytical Procedures, International Conference on Harmonization,  Geneva,October 1991, 1-5.

3.       ICH, Q2B, Validation of Analytical Procedures: Methodology, International Conference on Harmonization, Geneva, November, 1996, 1-8.

4.       Kamboj, P.C., Pharmaceutical Analysis, 1st Edn., Vallabh.Publications, New Delhi, 2003, 1, 155.

5.       United States Pharmacopoeia. XXIII, 22nd Revision, United States of Pharmacopoeia Convention, Inc., U.S.A, 1995, 2132-2136.

6.       Gupta, S.C., Fundamentals of Statistics, 4th Edn., Himalaya publishing house,New Delhi , 1999


 

 

 

 

Received on 25.04.2009       Modified on 23.06.2009

Accepted on 21.07.2009      © RJPT All right reserved

Research J. Pharm. and Tech. 3(2): April- June 2010; Page 409-411