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Harshil R. Patel*, Sejal K. Patel
Department of Quality Assurance,
S. K. Patel College of Pharmaceutical Education and Research,
Ganpat University, Ganpat Vidyanagar – 384012, Mehsana, Gujarat, India.

The present manuscript describes simple, sensitive, rapid, accurate, precise and economical derivative spectroscopic methodfor the simultaneous determination of Eperisone Hydrochloride(EPE) and Lornoxicam (LOR) in synthetic mixture. Derivative spectroscopy offers a useful approach for the analysis of drugs in mixtures. In this study a first-derivative spectroscopic method was used for simultaneous determination of Eperisone Hydrochloride and Lornoxicam using the zero-crossing technique. The measurements were carried out at wavelengths of 264 nm and 225.2 nm for Eperisone Hydrochloride and Lornoxicam  respectively. The method was found to be linear (r2>0.998) in the range of 2- 30 μg/ml for Eperisone Hydrochloride at 264 nm. The linear correlation was obtained (r2>0.996) in the range of 2-14 μg/ml for Lornoxicam at 225.2 nm. The limit of detection was 0.2565 and 0.235 μg/ml for Eperisone Hydrochloride and Lornoxicam respectively. The limit of quantification was 0.7774 and 0.7121 μg/ml respectively. The method was successfully applied for simultaneous determination of Eperisone Hydrochloride and Lornoxicam in synthetic mixture.


Eperisone Hydrochloride (EPE) is a well known antispasmodic drug1. Chemically it is 4-ethyl-2-methyl-3-piperidinopropiophenone hydrochloride (Figure 1). It is official in Japanese Pharmacopoeia (JP)2. JP describes potentiometric titration method for its estimation. Literature survey reveals liquid chromatography-ESI- tandem mass spectrometry for determination of eperisone in human plasma3. Literature survey also reveals Liquid Chromatography – Electrospray Ionization - Mass Spectrometry4, GC-MS5 method for the determination of Eperisone in human plasma. Lornoxicam (LOR) (3E)-6-chloro-3-[hydroxy (pyridin-2- ylamino)  methylene]-2-methyl-2,  3-dihydro-4H-thieno  [2,3-e]  [1,  2]  thiazin-4-one  1,  1-dioxide (Figure 2) is  a  novel  non- steroidal  anti-inflammatory  drug  (NSAID)  with  marked analgesic activity6. Various analytical methods, such as RPHPLC7, Extractionless HPLC8, UV spectroscopy9, HPTLC10, LC-MS-MS11  determination of Lornoxicam in dosage forms and human plasma. Lornoxicam in combination with other drug like Diacerin12 and Thiocolchicoside13 have been also detected. The combination of these two drugs is not official in any pharmacopoeia; hence no official method is available for the simultaneous estimation of EPE and LOR in their combined synthetic mixture or dosage forms. Literature survey does not reveal any simple spectrophotometric method for simultaneous estimation of EPE and LOR in synthetic mixture or combined dosage forms .The present communication describes simple, sensitive, rapid, accurate, precise and cost effective spectrophotometric method based on first order derivative for simultaneous estimation of both drugs in synthetic mixture.

Figure 1: Chemical structure of Eperisone hydrochloride (EPE)

Figure 2: Chemical structure of Lornoxicam (LOR)


A Shimadzu model 1700 (Japan) double beam UV/Visible spectrophotometer with spectral width of 2 nm, wavelength accuracy of 0.5 nm and a pair of 10 mm matched quartz cell was used to measure absorbance of all the solutions. Spectra were automatically obtained by UV-Probe system software. A Sartorius CP224S analytical balance (Gottingen, Germany), an ultrasonic bath (Frontline FS 4, Mumbai, India) was used in the study.

Reagents and materials
EPE bulk powder was kindly gifted by Sun Pharmaceuticals Ltd., Vadodara, Gujarat, India and LOR bulk powder was kindly gifted by Acme Pharmaceuticals Ltd., Mehsana, Gujarat, India. Methanol (AR Grade, S. D. Fine Chemicals Ltd., Mumbai, India) and Whatman filter paper no. 41 (Millipore, USA) were used in the study.

Preparation of standard stock solutions
An accurately weighed standard EPE and LOR powder (10 mg) were weighed and transferred to 100 ml separate volumetric flasks and dissolved in methanol. The flasks were shaken and volumes were made up to mark with methanol to give a solution containing 100 μg/ml of each EPE and LOR.

Determination of the zero crossing points
This method is based on first order derivative spectroscopy to overcome spectral interference from other drug. Zero order spectrums of both the drugs were converted to first order derivative spectra with the help of spectra manager software.

It was observed that LOR showed dA/dλ zero at 264 nm in contrast to EPE that has considerable dA/dλ at this wavelength. Further, EPE has zero dA/dλ at 225.2 nm while at this wavelength LOR has significant dA/dλ. Therefore wavelengths 264 nm and 225.2 nm were employed for the determination of EPE and LOR respectively without interference of other drug. The calibration curves were plotted at these two wavelengths of concentrations against dA/dλ separately. Seven working standard solutions having concentration 2, 5, 10, 15, 20, 25 and 30 μg/ml for EPE and 2, 4, 6, 8, 10, 12 and 14 μg/ml for LOR were prepared in methanol and the absorbances at 256 nm (zero crossing point for LOR) and 225.2 nm (zero crossing point for EPE) were measured and the calibration curves were plotted at these wavelengths.

Validation of the proposed method
The proposed method was validated according to the International Conference on Harmonization (ICH) guidelines14.

Linearity (Calibration curve)
The calibration curves were plotted over a concentration range of 2-30 μg/ml for EPE and 2-14 μg/ml for LOR. Accurately measured standard solutions of EPE (0.2, 0.5, 1.0, 1.5, 2.0, 2.5, 3.0ml) along with LOR (0.2, 0.4, 0.6, 0.8, 1.0, 1.2, 1.4ml) were transferred to a series of 10 ml of volumetric flasks and diluted to the mark with methanol. The absorbances of the derivatised spectra were measured at 264 nm and 225.2 nm for EPE and LOR respectively against methanol as blank. Six replicate analysis were carried out. Absorbance Vs concentration were plotted to obtain the calibration graph. Both drugs obey the Beer‘s law with the above concentration range with R2 value of 0.998 and 0.996 for EPE and LOR, respectively.

Method precision (Repeatability)
The precision of the instrument was checked by repeated scanning and measurement of absorbance of solutions (n = 6) for EPE (10 µg/ml) and LOR (6 µg/ml) without changing the parameter of the proposed spectrophotometry method.

Intermediate precision (Reproducibility)
The intraday and interday precision of the proposed method was determined by analyzing the corresponding responses 3 times on the same day and on 3 different days over a period of 1 week for 3 different concentrations of standard solutions of EPE and LOR (5, 15, 25 µg/ml for EPE and 4, 8, 12 µg/ml for LOR). The result was reported in terms of relative standard deviation (% RSD).

Accuracy (Recovery study)
The accuracy of the method was determined by calculating recovery of EPE and LOR by the standard addition method. Known amounts of standard solutions of EPE and LOR were added at 75, 100 and 125 % level to prequantified sample solutions of EPE and LOR (10 µg/ml for EPE and 6 µg/ml for LOR). The solutions were measured at 264 nm for EPE and 225.2 nm for LOR and % recovery of the sample were calculated. The experiment was repeated for three times.



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