- *Corresponding Author:
- I. Singhvi

Department of pharmaceutical sciences, M. L. sukhadia university, Udaipur - 313 001, India

**E-mail:**[email protected]

Date of Submission | 19 August 2006 |

Date of Revision | 4 September 2007 |

Date of Acceptance | 28 November 2007 |

Indian J Pharm Sci, 2007, 69 (6): 780-783 |

## Abstract

Three simple, accurate, economical and reproducible spectrophotometric methods for simultaneous estimation of two-component drug mixture of rosiglitazone maleate and glimepiride in combined tablet dosage form have been developed. Developed methods are based on direct estimation of rosiglitazone maleate at 318.0 nm, as at this wavelength glimepiride has zero absorbance and hence does not interfere. For estimation of glimepiride first developed method involves formation and solving of simultaneous equation at 238.0 nm. Second developed method makes use two wavelength spectroscopy using 244.8 nm and 257.2 nm as two wavelengths. Third developed method is based on first derivative spectroscopy using 252.0 nm as zero crossing point for estimation of glimepiride. All the developed methods obey Beer's law in the concentration ranges employed for the respective methods. The results of analysis were validated statistically and by recovery studies.

## Keywords

Simultaneous spectrophotometric analysis, Rosiglitazone maleate, Glimepiride

Rosiglitazone maleate (RSGN) is a new oral antidiabetic drug and chemically, it is (±)-5-{p- [2- (methyl-2-pyridylamino)ethoxy]benzyl}-2,4- thiazolidinedione maleate [1]. The drug is not yet official in any of the pharmacopoeia. For analysis of RSGN two LC [2,3], one MEKC (Micellar Electrokinetic Chromatographic) [4] and three HPLC [5-7] methods have been reported in literature. Glimepiride (GLIM), is a sulphonylurea antidiabetic drug and chemically it is 3-ethyl-2,5-dihydro-4-methyl-N-[2-[4-[[[[(trans-4- methylcyclohexyl)amino]carbonyl]amino]sulphonyl] phenyl]ethyl] 2-oxo-1H-pyrrole-1-carboxamide [8]. The drug is not yet official in any of the pharmacopoeia. Literature survey revealed one spectrophotometric [9] and three HPLC [10-12] methods have been reported for the estimation of GLIM in pharmaceutical formulations. Although, some methods have been reported for the estimation of RSGN and GLIM from single component formulation, there is no single method reported for simultaneous estimation of these two drugs from combined tablet dosage form. An attempt in the present study has been made to develop simple, accurate and economical method for simultaneous estimation of RSGN and GLIM from combined tablet dosage form. The result of analysis using the developed spectrophotometric methods for simultaneous estimation was found to be satisfactory such that the developed methods can be used for routine analysis of drugs from combined pharmaceutical dosage form.

## Materials and Methods

A PC based Systronic, UV/Vis double beam spectrophotometer (model No. 2101) with spectral bandwidth of 2 nm, wavelength accuracy ±0.5 nm (with automatic wavelength correction) and wavelength readability 0.1 nm increment was employed for all measurements using a matched pair of 10 mm quartz cells.

Standard bulk drug samples of RSGN and GLIM were provided by Dr. Reddy’s Laboratories, Hyderabad. Sodium hydroxide solution (0.1N) was used as solvent for the preparation of stock solution and for further dilutions. Double distilled water was used for the preparation of 0.1 N sodium hydroxide solution. The tablet samples of combined dosage form of RSGN and GLIM (Rosicon G, Glenmark Pharmaceuticals, Mumbai and Enseline-2G, Torrent Pharmaceuticals Ltd., Ahmedabad) were procured from local pharmacy.

**Method I, simultaneous equation method**

From the overlain spectra of RSGN and GLIM in 0.1
N sodium hydroxide solution (**fig. 1**), it was observed
that GLIM has zero absorbance at 318.0 nm, where
RSGN has substantial absorbance so RSGN was
estimated directly at this wavelength. For estimation
of GLIM simultaneous equation was framed. Pure
drug sample of RSGN and GLIM were dissolved
separately in 0.1 N sodium hydroxide solution so as
to give several dilutions of standard in concentration
range of 0-30 µg/ml of each drug. All dilutions were
scanned in wavelength range of 210.0 nm-370.0 nm.
Absorbance was measured at 318.0 nm against reagent
blank and plotted a calibration curve. For estimation
of GLIM simultaneous equation was framed on the
basis of absorptivity coefficient of two drugs at 238.0
nm. Absorptivity coefficient for GLIM at 238.0 nm
was calculated as 418.63 cm^{-1}g^{-1}l and for RSGN
394.72 cm^{-1}g^{-1}l. These calculated values are the means
of four independent determinations. Equation framed
for GLIM estimation was, A= 418.63 C_{1}+394.72 C_{2},
where A is absorbance of sample solution at 238.0
nm, C_{1} and C_{2} are concentration of GLIM and RSGN
respectively in g/l. Validity of above framed equation
was checked by preparing five mixed standards using
pure drug sample of two drugs, measuring their
absorbance at respective wavelengths and calculating
concentration of two components. Results of which
are reported in **Table 1**.

Sample no. | Conc. Present | %Conc. Found | ||||
---|---|---|---|---|---|---|

µg/ml | Method I | Method III | ||||

RSGN | GLIM | RSGN | GLIM | RSGN | GLIM | |

1 | 5 | 30 | 100. | 99. | 100. | 102. |

2 | 10 | 25 | 99. | 100. | 99. | 103. |

3 | 15 | 20 | 98. | 99.10 | 98. | 103. |

4 | 20 | 15 | 101. | 100. | 101. | 102. |

5 | 25 | 10 | 100. | 101. | 100. | 100.00 |

RSGN is rosiglitazone maleate, GLIM is glimepiride

**Table 1:** Results of validation studies for method I and Iii using mixed standards

**Method II, two wavelength calculation method**

Estimation of RSGN was carried out in a manner
similar to method-I. In this developed method for
estimation of GLIM two wavelength calculation
method was used. Two wavelengths selected for estimation of GLIM are 244.8 nm and 257.2 nm
based on the principle that absorbance difference
between two points on a mixture spectra is directly
proportional to concentration of component of interest
and independent of interfering component. Five mixed
standards of pure drugs containing 0-30 µg/ml of
each drug were prepared in 0.1N sodium hydroxide
solution. Absorbance of all standards were recorded at
two wavelengths 244.8 nm and 257.2 nm, determined
absorbance difference (A_{1}-A_{2}) values and plotted
calibration curve between absorbance difference values
and concentration of drug for estimation of GLIM.

**Method III, first order derivative spectroscopy method**

Estimation of RSGN was carried out in a manner
similar to method-I. For estimation of GLIM first
order derivative spectroscopy method was used. From
first order derivative spectra of RSGN and GLIM in
0.1 N sodium hydroxide (**fig. 2**) zero crossing point
252.0 nm was selected for estimation of GLIM.
Accurately weighed pure drug sample of RSGN and
GLIM were dissolved in 0.1 N sodium hydroxide
solution so as to give several dilutions in range of
0-30 µg/ml of each drug. The absorbance of these
dilutions was recorded in first order derivative mode
at 252.0 nm for estimation of GLIM and calibration
curve was prepared. Validity of proposed method
was checked by preparing five mixed standards using pure drug sample of two drugs and absorbance
was measured at selected zero crossing point and
determined concentration of GLIM using calibration
curve. Results of validation studies are reported in
**Table 1**.

**Analysis of commercial formulation**

For method I, twenty tablets of each brand were
weighed accurately and finely powdered. From each
triturate of the twenty tablets an amount equivalent to
2 mg rosiglitazone maleate was accurately weighed
and extracted four times with 20 ml portions of 0.1
N sodium hydroxide solution and filtered through
Whatman filter paper no. 41 into a 100 ml volumetric
flask. Washed residue with 0.1N sodium hydroxide
solution and added washings to filtrate, volume of
filtrate was made to 100 ml mark with 0.1N sodium
hydroxide solution. Absorbance of this dilution was
measured at 238.0 nm and 318.0 nm. Concentration of
RSGN was calculated directly from absorbance value
at 318.0 nm from calibration curve prepared using
standard drug solution. Concentration of GLIM was
calculated using above framed equation. Results of
analysis of tablet formulation are reported in **Table 2**.

Method | Batch | Label claim mg/Tab | % Label claim estimated* | Standard deviation | % Conc. Recovered** | ||||
---|---|---|---|---|---|---|---|---|---|

RSGN | GLIM | RSGN | GLIM | RSGN | GLIM | RSGN | GLIM | ||

Method I | A | 2 | 1 | 100. | 101. | 0.4171 | 0.3040 | 100. | 100. |

B | 2 | 1 | 100. | 101. | 0.8343 | 0.0848 | 100. | 99. | |

Method II | A | 2 | 1 | 100. | 101. | 0.4171 | 0.3889 | 100. | 100.30 |

B | 2 | 1 | 100. | 101.10 | 0.8343 | 0.7778 | 101. | 101. | |

Method III | A | 2 | 1 | 100. | 99. | 0.4171 | 0.7495 | 100. | 101. |

B | 2 | 1 | 100. | 101. | 0.8343 | 0.8697 | 100. | 98. |

*Averageofthreeestimations, **Averageofrecoverystudiesatthreedifferentconcentrationlevels.AisRosiconG(GlenmarkPharmaceuticals, Mumbai)andBisEnseline-2G (TorrentPharmaceuticalsLtd.,Ahmedabad). RSGNisrosiglitazone maleate,GLIMisglimepiride. MethodIisSimultaneousEquation method,MethodII isTwoWavelength Calculationmethod, MethodIII is Firstorder derivative spectroscopymethod

**Table 2:** Results of analysis of commercial formulation

For method II, preparation of tablet sample solution
and estimation of RSGN was carried out in a manner
similar to method I. For estimation of GLIM final
dilution was analyzed by recording absorbance at
244.8 nm and 257.2 nm and absorbance difference
values were noted and concentration was calculated
from the respective calibration curve. Results of
analysis are reported in **Table 2**.

For method III, preparation of tablet sample solution
and estimation of RSGN was carried out in a
manner similar to method I. For estimation of GLIM
absorbance of sample was recorded at 252.0 nm from
first order derivative spectra of sample and amount
of GLIM was calculated using respective calibration curve. Results of analysis are reported in **Table 2**.

**Recovery studies**

To study the accuracy, reproducibility and precision
for all the three developed methods, recovery studies
were carried out by the addition of standard drug
solution to pre-analyzed tablet sample with proper
dilutions at three different concentration levels with
in the range of linearity for both the drugs. Results of
recovery studies were found to be satisfactory and are
reported in **Table 2**.

## Results and Discussion

The proposed methods described for the simultaneous analysis of RSGN and GLIM in combined tablet dosage form has been found to be simple, accurate, rapid, economical and sensitive to be applied in routine analysis of tablets. In the described methods there are no additional extraction or separation procedures to extract the drug from the formulation excipient matrix thereby decreasing the error in quantitation.

The developed methods are based on direct estimation of RSGN at wavelength maxima of RSGN, where GLIM has no absorbance, however for estimation of GLIM special techniques were used. First developed method involving formation and solving of simultaneous equation is based on absorptivity coefficient of two drugs at wavelength maxima of GLIM i.e. once the equation is framed then it is just required to measure the absorbance of sample solution at selected wavelengths and few calculations that can be manually done. Framed equation was validated using laboratory prepared mixed standards of two drugs which gave satisfactory results.

Second developed method for estimation of GLIM makes use of two wavelength calculation method so as to remove interference between two components. Proper selection of two wavelengths for estimation of a component is critical.

Third developed method for simultaneous analysis of RSGN and GLIM from combined dosage form makes use of first order derivative ultraviolet spectrophotometry based on principle that at zero crossing point of one component, the other component has substantial absorbance.

The results of analysis of two drugs from tablet formulation using all the three developed methods were found close to 100% for both RSGN and GLIM, standard deviation was satisfactorily low indicating accuracy and reproducibility of the methods. Recovery studies were satisfactory which shows that there is no interference of excipients. The developed methods were found to be simple, rapid, accurate and can be used for routine estimation of two drugs from tablet formulations.

## Acknowledgements

Authors would like to acknowledge Dr. Reddy’s Laboratories, Hyderabad, India for providing pure drug samples of RSGN and GLIM.

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