INTRODUCTION:

Domperidone is an antiemetic and antinauseant and acts on dopamine receptor system as an antagonist. Chemically it is 5-chloro-1-[1-(2,3-dihydro-2-oxo-1H-benzimidazole-1-yl)propyl)-4-piperidyl]-2,3-dihydro-1H-benzimidazol-2-one1. It is official in British Pharmacopoeia and European Pharmacopoeia where non-aqueous titration is the official method of assay^.1-3

Omeprazole is 5-methoxy-2-(4-methoxy-3,5-dimethyl-2pyridinylmethylsulfinyl)-1H-benzimidazole. It is official in IP 2,USP 3 and BP. Omeprazole is the proton pump inhibitor. In the acidic conditions of the stomach, omeprazole react with cysteine group in H+/K+- ATPase, thereby destroying the ability of the parietal cells to produce gastric acid. Thus together both these drugs have synergistic effect in controlling the gastric ulcer diseases^.4-7

Literature survey reveals that several methods like Spectrophotometry,HPLC,HPTLC and LC-MS were reported for the determination of Domperidone in combination with other drugs as well as in biological fluids but no method has been reported for simultaneous analysis of Domperidone and Omeprazole in its combination. anywhere before. These above developed methods are too expensive and time consuming.

An attempt has been made to develop a simple, economical,precise ,accurate and reproducible spectrophotometric method for estimation of Domperidone and Omeprazole in bulk as well as pharmaceutical formulations.^8-10

MATERIALS AND METHODS:

REAGENTS AND CHEMICALS:

Pure Domperidone was obtained from Macro Lab (Bangalore) and pure omeprazole was obtained from Lupine Research Pvt. Limited, Pune. The commercial fixed dose capsule formulation OMEZ-D(DR.REDDY’S) containing 10 mg Domperidone were procured from the local market. Dimethyl Formamaide reagents used were of AR grade.Distilled water was used during the experiment. Spectral and absorbance measurements were made on Analytical technologies spectrophotometer with 1 cm matched quartz cells.

INSTRUMENTATION:

A Shimadzu UV/Visible spectrophotometer, model 1700 (Japan) was employed with spectral bandwidth of 2 nm and wavelength accuracy of ± 0.5 nm, with automatic wavelength correction employing a pair of quartz cells. A Shimadzu electronic analytical balance (AX-200) was used for weighing the sample.

PREPARATION OF STANDARED STOCK SOLUTION:

A. Standard Domperidone stock solution (100 µg/mL):

Domperidone standard stock solution was prepared by weighing 10 mg of Domperidone, transferred to a 100 ml volumetric flask and volume was made up to 100 ml with Dimethyl Formamide and Water in the ratio of 20: 80(Dimethyl Formamide : Water) to get a concentration of 100μg/ml.The prepared solution is sonicated for 5 minutes and filtered through the whatman filter patae no. 41.^12-14

B. Standard Omeprazole stock solution (100 µg/mL):

Omeprazole standard solution was prepared by weighing 10 mg of Omeprazole and transferred to a 100 ml volumetric flask and volume was made up to 100 ml with with Dimethyl Formamide and Water in the ratio of 20: 80 to get a concentration of 100 μg/ml. The prepared solution is sonicated for 5 minutes and filtered through the whatman filter patae no. 41.^14-16

CALIBRATION CURVE:

A calibration curve was plotted over a concentration range of 5-30 mg/mL for omeprazole and 18-28 for Domperidone. Accurately measured standard stock solution of Domperidone (1.8, 2, 2.2, 2.4, 2.6 and 2.8mL) and standard stock solution of Omeprazole (0.5, 1, 1.5, 2, 2.5 and 3mL) were transferred to a separate series of 10 mL of volumetric flasks and diluted to the mark with Dimethyl Formamide. The absorbance of each solution was measured at both the wavelengths 290 nm and 301nm.

FIGURE NO.1-CALIBRATION CURVE OF DOMPERIDONE

FIGURE NO.2-CALIBRATION CURVE OF DOMPERIDONE AT ISOBESTIC POINT

Calibration curves were constructed for Domperidone and Omeprazoleby plotting absorbance versus concentrations at both wavelengths. Each reading was average of five determinations.^17-18

FIGURE NO.3-CALIBRATION CURVE OF OMEPRAZOLE

FIGURE NO.4-CALIBRATION CURVE OF OMEPRAZOLE AT ISOBESTIC POINT

SELECTION OF ANALYTICAL WAVELENTH:

For selection of analytical wavelength for the Q-absorbance method (Method-1)

The stock solutions of Domperidone and Omeprazole were separately diluted in Dimethyl Formamide and water in the ratio of 20:80 to get a concentration of 10 μg/ml of Domperidone and 10 μg/ml of Omeprazole respectively and scanned in the wavelength range of 200 -400 nm. From the overlain spectra of both drugs wavelengths 290.9 nm (isoabsorptive point), 284.8 nm (λ max of Domperidone) and 301.2 nm(λ max of Omeprazole) were selected for the formation of Q-absorbance equation. The absorbance of various dilutions of Domperidone measured at 284.8 nm and calibration curves were plotted. Similarly the absorbance of various dilutions of Omeprazole measured at 301.2 nm, calibration curves were plotted. The absorptivities (A1%, 1cm) of each drug at both the wavelengths were also determined. The absorbance and absorptivity values at the particular wavelengths were calculated and substituted in the following equation, to obtain the concentration.^18-19

FIGURE NO.5- SPECTRUM OF OMEPRAZOLE

FIGURE NO.6- SPECTRUM OF DOMPERIDONE

FIGURE NO.7- SPECTRUM OF OVERLAY OF OMEPRAZOLE AND DOMPERIDONE

METHODS:

A) ABSORPTION RATIO/Q METHOD ANALYSIS:²⁰²¹

From the over line spectrum of Domperidone and Omeprazole, one wavelength was selected for the estimation of both drugs, which is known as isoabsorptive point (at 290.9nm). The dilutions of standard and sample solutions were prepared. The absorptivity values were determined at 290.9nm.The method employs Q values and the concentrations of drugs in sample solution were determined by using the following formula

C_DOM = (QM – QY) X A1 / (QX – QY) X ax1,

C_OME= ( QM - QX) X A1/ (QY – QX) X ax2,

Where,

C_DOM= concentration of Domperidone ,

C_OME = concentration of Omeprazole respectively,

A1 = absorbance of sample at 290.9 nm,

For Domperidone-

ax1 = the absorptivity of Domperidoneat 290.9nm

QX, QY and QM was obtained using the following equation

QX = (absorptivity of Domperidone at 284.8 nm)

(absorptivity of Domperidone at 290.9 nm)

QY = (absorptivity of Omeprazole at 284.8 nm)

(absorptivity of Omeprazole at 290.9 nm ) and

QM = (absorbance of sample at 284.8 nm)

(absorbance of sample at 290.9 nm).

For Omeprazole-

ax2 = the absorptivity of Omeprazoleat 290.9 nm

nm, QX,QY and QM was obtained using the following equation

QX = (absorptivity of Omeprazole at 301.2 nm)

(absorptivity of Omeprazole at 290.9 nm),

QY = (absorptivity of Domperidone at 301.2 nm)

(absorptivity of Domperidone at 290.9 nm ) and

QM = (absorbance of sample at 301.2 nm)

(absorbance of sample at 290.9 nm).

B) SIMULTANEOUS ESTIMATION METHOD:

The spectra of Domperidone and Omeprazole of method 1 was used and wavelength 284.8 and 301.2 nm (λ max of DOM and λ max of OME) were selected for the formation of the simultaneous equations. For calibration curves, stock solutions of Domperidone and Omeprazole in the concentration of range of 18 – 28 μg/ml and 5 – 30 μg/ml respectively. The absorbance of Domperidone and Omeprazole were measured at 284.8 and 301.2 nm, calibration curves were plotted. The absoptivities of both the drugs at both the wavelengths were determined (Table-1).^22-23

TABLE NO. 1-OPTICAL CHARACTERISTICS DATA

Parameter	Method І Q-Absorbance Ratio Method		Method II Simultaneous Equation Method
	DOM	OME	DOM	OME
Working λmax	284.8 and 290.9	301.2 and 290.9	284.8	301.2
Beer’s Low Limit	18-28μg/ml	5-30 μg/ml	18-28 μg/ml	5-30 μg/ml
Correlation coefficient*	0.9998	0.9990	0.9997	0.9997
Intercept*	-0.0263	0.0353	-0.0045	0.0333
Slope*	0.0225	0.0250	0.0306	0.0324
Molar Absorptivity(lit/mol/cm)	9582.75	8635.5	13032.54	11191.61
Regression Equation	Y=0.0225x-0.0263	Y=0.0250+ 0.0250	y=0.0306x-0.0045	Y=0.0324x+ 0.033

* Average of six determination; DOM=Domperidone; OME=Omeprazole.

TABLE NO. 2-RESULT OF CAPSULE ANALYSIS

Method	Drug Name	Lable Claim in mg	% Lable Claim Found*	*Amount Found in mg**
І	DOM	30 mg	99.06 %	29.72 mg
	OME	20 mg	95.53%	19.11mg
ІІ	DOM	30 mg	96.00%	28.80 mg
	OME	20 mg	78.47%	15.71 mg

DOM- Domperidone; OME-Omeprazole; *Average of six estimation of Capsule formulation.

The absorbance and the absorptivity values at the particular wavelength were calculated and substituted in the following equation, to obtain the concentration.

C_DOM = (A1ax2 – A2ax1) / (ax2ay1 – ax1ay2).

C_OME = (A2ay1 – A1ay2) / (ax2ay1 – ax1ay2).

Where,

C_DOM= Concentration of Domperidone

C_OME = Concentration of Omeprazole respectively,

A 1 = absorbance of sample at 284.8 nm,

A 2 = absorbance of sample at 301.2 nm,

ax1 = absorptivity of Domperidone at 284.8 nm and

ax2 = absorptivityof Domperidone at 301.2 nm,

ay1 = absorptivity of Omeprazole at 284.8 nm and

ay2 = absorptivity of Omeprazole at 301.2 nm,

ANALYSIS OF FORMULATION:

Twenty Capsules of brand OMEZ D (SR) (DR. REDDY’S) containing 20 mg of Omeprazole and 30 mg of Domperidone were weighed, average weight determined and finely powdered with the help of mortor and pestle. Appropriate quantity of powder from each tablet equivalent to 30 mg of Dmperidone was accurately weighed transferred to a 100 ml volumetric flask and volume was made up to 100 ml with Dimetyl Formamide and water in the proportion of 20:80. shaken vigorously for 15 mins and filtered through the whatman filter paper no.41. Necessary dilutions of filtrate were made with Dimethyl Formamide and water to get final concentration 15 μg/ml of Omeprazoleand 15 μg/ml of Domperidone. Absorbance of this solution was measured at 301.2 nm (λmax of Omeprazole) 284.8nm((λmax of Domperidone), and 290.9 nm (Isobestic Point), values were substituted in the respective formulae of (Method 1 and 2) to obtain concentration .Results are shown in the following Table-2.^23-25

VALIDATION:^24-26

Validation of the developed method was done according to the USP 2006, Asian edition

LINEARITY:

The linearity of the method is its ability to elicit test results that are directly proportional to the concentration of the analyte in samples. The calibration curve was taken in the range of 18-28μg/ml for Domperidone and 5-30 μg/mL for Omeprazole at the respective λmax. The correlation coefficient of the linearity was found to be 0.999 at each wavelength for both drugs as shown in table 1.

RECOVERY STUDY:

In order to ensure the reliability and suitability of the proposed method, recovery studies were carried out. It was done by mixing known quantity of standard drug with formulation sample and the content were reanalysed by the proposed method. To a quantity of formulation equivalent to 30 mg of Domperidone, standard drugs of Domperidone and Omeprazole were added at 80%, 100% and 120% levels. This was extracted, diluted and reanalysed as per the formulation procedure. Absorbance were noted at respective wavelength. Recovery studies were repeated for six times and the results are shown in the table-3.

TABLE NO. 3-RECOVERY STUDY OF OMEPRAZOLE AND DOMPERIDONE

Method

Recovery Level

% Recovery

S.D

% RSD OR %COV

% Recovery

S.D

% RSD OR %COV

80 %

100%

120%

Domperidone

Omeprazole

98.25

99.22

99.60

0.05049

0.07416

0.06557

0.64246

0.83048

0.65837

100.61

99.51

98.50

0.15524

0.13304

0.0300

1.928

1.4848

0.003045

ІІ

80%

100%

120%

97.62

97.78

98.80

0.05567

0.02645

0.04582

0.7129

0.2986

0.46382

100

99.8

0.06519

0.0600

0.0300

0.8149

0.6734

0.3006

S.D-Standard deviation; RSD-Relative standard deviation; COV-Coefficient of variation

TABLE NO.4- PRECISION STUDY OF OMEPRAZOLE AND DOMPERIDONE

Day	Method І		Method ІІ
Interday	% of lable claim estimated ( Mean± % RSD)
	DOM	OME	DOM	OME
	103.12±0.00445	98.46±0.169	104.2±0.8093	97.43±0.4464

TABLE NO. 5- LOD AND LOQ RESULTS OF OMEPRAZOLE AND DOMPERIDONE

Validation Parameter	Method I (Q- Absorbance Ratio Method)		Method II (Simultaneous equation method)
	Domperidone	Omeprazole	Domperidone	Omeprazole
LOD(µg/ml)	0.44484	0.400356	0.28125	0.14400
LOQ(µg/ml)	1.18000	1.21000	0.85228	0.43442

LOD- Limit of detection; LOQ- Limit of quantification

PRECISION:

The precision of an analytical method is determined by assaying a sufficient number of aliquots of a homogeneous sample to be able to calculate statistically valid estimate of % Relative Standard Deviation (%RSD). Intermediate precision was done to express within laboratory variation, on different days. Five replicates of 15 μg/mL concentration of the working standard mixture and sample solution were analysed %RSD was found to be less than 2% (Table-4).

SPECIFICITY:

Results of tablet solution showed that there is no interference of the excipients when compared with the working standard solution. Thus, the method was said to be specific.

LIMIT OF DETECTION:

It is the lowest amount of analyte in a sample that can be detected but not necessarily quantitated under the stated experimental conditions. Limit of detection can be calculated using following equation as per ICH guidelines (Table-5).

LOD = 3.3 × N/S

Where,

N = Standard deviation of the response and

S = Slope of the corresponding calibration curve.

LIMIT OF QUANTIFICATION:

It is the lowest concentration of analyte in a sample that can be determined with the acceptable precision and accuracy under stated experimental conditions. Limit of quantification can be calculated using following equation as per ICH guidelines (Table-5).

LOQ = 10 × N/S

Where,

N = Standard deviation of the response and

S = Slope of the corresponding calibration curve

The overlain spectra of both the drugs showed that the peaks are well resolved, thus satisfying the criteria for obtaining maximum precision, based on absorbance ratio. The criteria being the ratios (A2/A1) / (ax2/ax1) and (a y2 / ay1) / (A2/A1) should lie outside the range 0.1 – 2.0 for precise determination of (Y) and (X) respectively. Where A 1/A2 represents the absorbance of mixture at λ1 and λ2, ax1 and ax2 denote absoptivities of (X) at λ1 and λ2 and ay1 and ay2 denote absoptivities of (Y) at λ1 and λ2 respectively. In this context, the above criterion was found to be satisfied for Domperidone(X) and Omeprazole(Y). Where λ1 (284.8 nm) and λ2 (290.9 nm) for Q-absorbance method, λ1 (284.8 nm) and λs2 (301.2 nm) for simultaneous equation method

RESULT AND DISCUSSION:

The proposed methods for simultaneous estimation of Domperidone and Omeprazole in combined capsule dosage form were found to be simple, accurate, economical and rapid. The % RSD was found to be less than 2% in the developed method. Hence proposed method may be used for routine analysis of these drugs in combined dosage forms.

ACKNOWLEDGEMENT:

The authors are thankful to Lupine Pharmaceuticals Pvt Ltd. and Macro Lab(Banglor)

for providing Omeprazole and Domperidone as gift samples for this work. And also Thanks to Prof., K.B.Burade, (Incharge Principal) GOVERNMENT COLLEGE OF PHARMACY, KARAD, for providing the required facilities for my research work.

I am also greetfull of Mrs.A.S.Kulkarni (Assistant Professor) for guiding me.

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Received on 30.04.2011 Modified on 12.05.2011

Asian J. Research Chem. 4(7): July, 2011; Page 1119-1124