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DEVELOPMENT AND VALIDATION OF RP-HPLC METHOD FOR SIMULTANEOUS ESTIMATION OF ILAPRAZOLE AND DOMPERIDONE IN PHARMACEUTICAL DOSAGE FORM

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ABOUT AUTHORS:
R.A Tamboli, V.C. Chauhan, M.M. Pathan, S.K. Tirgar, D.A. Shah, R.R. Parmar
APMC College of Pharmaceutical Education and Research,
Motipura, Himmatngar, Gujarat
tambolirushabh@gmail.com

ABSTRACT
A specific,  accurate,  precise  and  reproducible  RP-HPLC  method  has  been  developed  and subsequently validated for the simultaneous determination of Ilaprazole and Domperidone  in pharmaceutical dosage form. The proposed HPLC method utilizes hypersil (Thermo scientific) C18 column (250 mm × 4.6 mm id, 5 μm particle size), and mobile phase consisting of methanol:phosphate buffer (40:60) and pH adjusted to 4.0 with 0.1M glacial acetic acid at a flow rate of 1.0 mL/min. Quantitation was achieved with UV detection at 230 nm based on peak area with linear calibration curves at concentration ranges 5-15 μg/ml for Ilaprazole and 15-45 μg/ml for Domperidone. The retention time of  Ilaprazole and Domperidone were found to be 3.433 min and 5.860 min respectively. The method was validated in terms of accuracy,  precision,  linearity,  limits  of detection,  limits  of  quantitation  and  robustness. This method has been successively applied to marketed formulation and no interference from the formulation excipients was found.

REFERENCE ID: PHARMATUTOR-ART-2199

PharmaTutor (ISSN: 2347 - 7881)

Volume 2, Issue 7

Received On: 30/04/2014; Accepted On: 07/05/2014; Published On: 01/07/2014

How to cite this article: RA Tamboli, VC Chauhan, MM Pathan, SK Tirgar, DA Shah, RR Parmar; Development and Validation of RP-HPLC Method for Simultaneous Estimation of Ilaprazole and Domperidone in Pharmaceutical Dosage Form; PharmaTutor; 2014; 2(7); 149-156

INTRODUCTION
Ilaprazole is a proton pump inhibitor(PPI) used in the treatment of dyspepsia, Peptic ulcer disease (PUD), gastroesophageal reflux disease (GORD/GERD) and duodenal ulcer. It is available in strengths of 5, 10, and 20 mg. Clinical studies show that Ilaprazole is at least as potent a PPI as omeprazole when taken in equivalent doses. Studies also showed that Ilaprazole significantly prevented the development of reflux oesophagitis.[1].

Ilaprazole is chemically 2-[(RS)-[(4-methoxy-3-methylpyridin-2yl)methyl]sulfinyl]-5-(1H-pyrrol-1-yl)-1H-benzimidazole[3].

Fig.1.Chemical structure of Ilaprazole[4]

Domperidone chemically is 5-chloro-1-{1-[3-(2-oxo-2,3-dihydro-1H-1,3-benzodiazol-1-yl)propyl]piperidin-4-yl}-2,3-dihydro-1H-1,3-benzodiazol-2-one[7]. Fig.2

Domperidone acts as a gastrointestinal emptying (delayed) adjunct and peristaltic stimulant. The gastroprokinetic properties of domperidone are related to its peripheral dopamine receptor blocking properties. Domperidone facilitates gastric emptying and decreases small bowel transit time by increasing esophageal and gastric peristalsis and by lowering esophageal sphincter pressure. Antiemetic: The antiemetic properties of domperidone are related to its dopamine receptor blocking activity at both the chemoreceptor trigger zone and at the gastric level. It has strong affinities for the D2 and D3 dopamine receptors, which are found in the chemoreceptor trigger zone, located just outside the blood brain barrier, which - among others - regulates nausea and vomiting.[7]

Fig.2.Chemical structure of Domperidone[7]

Combination of Ilaprazole and Domperidone are used in treatment of peptic ulcer and gastroesophageal reflux disease (GORD/GERD) and duodenal ulcer.In the literature survey it was found that Ilaprazole and Domperidone were estimatedindividualy or in combination with other drugs by UV, HPLC, HPTLCSpectrofluorimethods[8-22]. But no method has been found for simultaneous estimation of Ilaprazole and Domperidone by chromatographic method. In the view of the need in the industry for routine analysis of Ilaprazole and Domperidone in formulation, attempts are being made to develop simple and accurate RP-HPLC method for simultaneous estimation of Ilaprazole and Domperidone and extend it for their determination in formulation.

MATERIAL AND METHOD
Equipment

RP–HPLC instrument equipped with SPD-20 AT UV-Visible detector, (LC-20AT, Shimadzu), Rheodyne injector (20 μl Capacity), BDS hypersil (Thermo scientific) C18 column (250 mm × 4.6 mm, 5 μ particle size) and Spinchrom software was used.

Chemicals and reagents
Reference standard of ILA and DOM were obtained from Montage laboratories PVT. LTD., Himatnagar. Methanol and used was of HPLC grade and Phophatebuffer(pH 4.0) and all other reagent were of AR grade.

Preparation of standard and test solutions
Preparation of mobile phase

Mobile phase were prepared by mixing of 400 ml of  methanol with 600 ml of phosphate buffer, whose pH was  adjusted to pH 4.0 by addition of glacial acetic acid. The mobile phase prepared was degassed by ultrasonication for 20 min, so as to avoid the disturbances caused by dissolved gases. The degassed mobile phase was filtered through 0.45 μ filters to avoid the column clogging due to smaller particles.

Preparation of standard stock solutions
An accurately weighed quantity of ILA(10 mg) and DOM(30 mg) were transferred to a 100 ml volumetric flask and dissolved and diluted to the mark with mobile phase to obtain standard solution having concentration of ILA (100 μg/ml) and DOM (300μg/ml).

Preparation of solutions for calibrationcurve
The calibration curves were plotted over the concentration range 5-15 μg/ml for ILA and 15-45μg/ml for DOM. From the stock solution 100 μg/ml of ILA and 300 μg/ml of  DOMprepared. From these working solutions of ILA and DOM (0.5 ml, 0.75 ml, 1.0 ml, 1.25 ml, 1.5 ml and 0.5 ml, 0.75 ml, 1.0 ml, 1.25 ml, 1.5 ml) were transferred to a series of 10 ml of volumetric flasks and diluted to the mark with mobile phase. Aliquots (20 μL) of each solution were injected under the operating chromatographic conditions described above.

Preparation of sample solution
Take quantity equivalent to 10 mg ILA and 30 mg DOM was transferred to 100 ml volumetric flask in Mobile Phase .The solution was filtered through whatman filter paper No. 41 and the volume was adjusted up to the mark with Mobile Phase. From this 1 ml of solution is diluted to 10 ml with the help of mobile phase to get final concentration of 10 μg/ml ILA and 30 μg/ml DOM.

METHOD VALIDATION[23-24]
The developed method was validated according to ICH guidelines. To check the system performance, the system suitability parameters were measured. System precision was determined on six replicate injections of standard preparations. Number of theoretical plates and asymmetry were measured.

Linearity
Linearity was performed with five concentrations ranging from 5-15μg/ml and 15-45 μg/ml for ILA and DOM respectively. The peak areas versus concentration of drug were plotted and a linear least-square regression analysis was conducted to determine the slope, intercept and correlation coefficient (r) to demonstrate the linearity of the method.

The limit of detection (LOD) and limit ofquantitation (LOQ)
LOD and LOQ of ILA and DOM were calculated using the following equations as per International Conference on Harmonization (ICH) guidelines.
LOD = 3.3 × σ/S
LOQ = 10 × σ/S

Where σ = the standard deviation of the response
S = Slope of calibration curve.

Precision
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 3 different concentrations of sample solutions of ILA (5μg/ml, 10 μg/ml and 15 μg/ml) and DOM (15 μg/ml, 30 μg/ml and 45 μg/ml). Percentage relative standard deviation (RSD) was calculated

Accuracy
Accuracy was performed by adding known amounts of ILA and DOM to the pre-analysed marketed formulation and then comparing the added concentration with the found concentration. Three levels of solutions were made which correspond to 80, 100 and 120% of the nominal analytical concentration (5 μg/ml for ILA and 15μg/ml for DOM). Each level was prepared in triplicate. The percentage recoveries of ILA and DOM at each level were determined. The mean recoveries and the relative standard deviation were then calculated.

Robustness
The robustness of the method was evaluated by assaying the test solutions after slight but deliberate changes in the analytical conditions i.e. flow rate (± 0.2 ml/ min), proportion of buffer and methanol (62:38 and 58:42 v/v), and pH of buffer (± 0.2).

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RESULT AND DISCUSSION
System Suitability

The chromatogram of ILA and DOM show retention time 3.473 min and 5.920 min respectively. Mobile phase used for separation was Phosphate buffer(pH 4.0) : Methanol (60:40) pH of buffer adjusted with glacial acetic acid. Standard chromatogram was given in Figure 3.System suitability parameters were shown in Table 1.

Figure 3: Standard Chromatogram of Ilaprazole and Domperidone

Table 1: System Suitability Parameters Of Chromatogram For ILA & DOM

PARAMETERS

ILA ± SD

(n = 6)

DOM ± SD

(n = 6)

Retention time (min)

3.473±0.022

5.920±0.016

Tailing factor

1.798±0.018

1.362±0.025

Theoretical plates

4910±2.62

6990±2.14

Resolution

10.163±0.98

Method validation
The calibration curves were plotted over the concentration range 5-15 μg/ml for ILA and 15-45 μg/ml for DOM are shown in figure 2 and figure 3 respectivaly. The data for intraday and interday precision for ILA and DOMare shown in Table 2 and Table 3respectively. Statistical analysis of recovery data is shown in Table 4. Results of robustness study of ILA and DOMare recorded in Table 5. It suggests that the developed method is robust. Summary of validation parameter is shown in Table 6.

Table 2: Linearity Data For ILA and DOM

ILA

DOM

Conc.μg/ml

Area* ± SD

Conc.μg/ml

Area* ± SD

5

665.892±13.62

15

850.283±10.56

7.5

983.147±13.92

22.5

1258.034±18.26

10

1344.146±15.26

30

1720.292±5.62

12.5

1554.965±18.2

37.5

2018.418±12.65

15

2013.587±30.25

45

2577.797±17.8

*Average of three determination

Figure 4: Caliberation curve of ILA Figure 5: Calibaration curve of DOM

Table 3: Intraday precision data for estimation of ILA and DOM

ILA

DOM

Conc.μg/ml

Peak Area* ± SD

%RSD

Conc.μg/ml

Peak Area* ± SD

%RSD

5

657.391±10.718

1.6304

15

 841.19±10.931

1.2997

10

1328.349±19.298

1.4528

30

1706.80±10.914

0.3195

15

1991.963±21.761

1.0924

45

2555.517±12.098

0.6749

Interday precision data for estimation of ILA and DOM

ILA

DOM

Conc.

μg/ml

Peak Area* ± SD

%RSD

Conc.

μg/ml

Peak Area* ± SD

%RSD

5

655.2037±13.31553

2.032273

15

839.377±10.80127

1.28682

10

1330.001±15.17277

1.140809

30

1708.712±5.46059

0.319573

15

1989.005±30.49116

1.532986

45

2556.956±17.2571

0.674908

Table 4: Recovery

Level of

recovery %

Amount of pure drug added (μg/ml)

HPLC Method % recovery

ILAPRAZOLE

DOMPERIDONE

ILAPRAZOLE

DOMPERIDONE

80

4

12

99.6689

100.3718

100

5

15

99.1231

99.5917

120

6

18

99.1576

100.0590

Mean % recovery

99.3165

100.0075

Standard Deviation

0.7059

0.6716

Relative Standard Deviation

0.7107

0.6706

Table 5: Robustness

Condition

                     Peak Area

ILA

DOM

Flow Rate

1.2 ml/ min

1303.544

1674.575

0.8 ml/ min

1383.888

1776.575

Mobile phase ratio

62:38

1302.376

1669.903

58:42

1370.602

1756.954

Ph

4.2

1275.916

1639.688

4.0

1369.845

1760.354

Average

1334.364

1713.008

S.D

23.082

15.469

% RSD

1.731

0.903

Assay of the tablet formulation
The proposed validated method was successfully applied to determine ILA and DOM in their capsule formulation shown in figure 6. The result obtained for ILA and DOM were comparable with the corresponding labelled amounts Table 4. No interference of the excipients with the peak of interest appeared.

Figure 6: Chromatogram of sample solution of ILA and DOM at 230 nm

Table 6: Assay of formulation

Capsule

mg/capsule

Assay(content in mg)*

(% of labelclaim*) ± % RSD

LUPILA-D

ILA

10

ILA

9.92

ILA

99.2± 0.80

DOM

30

DOM

30.42

DOM

101.4± 0.67

Table 7: Summary of Validation Parametere

PARAMETERS

RP-HPLC method

ILA

DOM

Concentration range (μg/ml)

5-15

15-45

Slope

130.69

56.205

Intercept

5.4642

-1.2

Correlation coefficient

0.9993

0.9995

LOD(μg/ml)

0.4826

0.9689

LOQ(μg/ml)

0.8659

1.1658

Repeatability (% RSD, n

= 6)

1.4858

0.5872

Precision (%RSD)

Interday (n = 3)

Intraday (n = 3)

1.5686

1.3956

0.7604

0.7647

Accuracy (% recovery),

(n=3)

99.3165±0.7059

100.0075±0.6716

% Assay

99.2± 0.80

101.4± 0.67

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CONCLUSION
The RP-HPLC method developed for analysis of Ilaprazole and Domperidonein their capsule dosage form is precise, accurate and with short run time. The method was fully validated showing satisfactory data for all the method validation parameters tested. The developed method is suitable for the quality control of the raw material, formulation and dissolution studies.

ACKNOWLEDGEMENT
The authors are grateful to Montage labs pvt. Ltd., Himatnagar for providing free gift sample of Ilaprazole and Domperidone. And authors are grateful to APMC college of pharmaceutical research and education for providing facility to carry out this work.

REFRENCES
1.KD Tripathi: Essentials of Medical pharmacology. Japee Brothers,2004
2.Moffat AC; Osselton MD; Widdop B and Watts J. Clarke’s Analysis of Drugs and poisons in pharmaceuticals.London Pharmaceutical press, 2011.
3.Indian Pharmacopoeia, Volume-III, 6thEdn, The Indian Pharmacopoeia commission, Ghaziabad, Govt. of India, Ministry of Health and Family Welfare, 2010.
4.Drug Bank, “Ilaprazole ”, drugbank.ca/drugs/DB00384.
5.Joel G.H, Perry B.M, Lee E.L, Raymond W.R, Alfred Cg, editor., Goodman Gilman’s The pharmacological basis of Therapeutics, 9th ed. New Jersy: Mc-Graw Hill Companies, 1996.
6.Sweetman SC. Martindale: The complete drug reference. London Pharmaceutical Press, 2009.
7.Drug Bank, “Domperidone”, drugbank.ca/drugs/DB00562.
8.Mansuri BK, Faldu S and Dadhania K. "Estimation of Ilaprazole in bulk drug and its pharmaceutical dosage form by difference spectrophotometric method".
9.Adhiyar P, Bhati S, Patel H and Pancholi S. "Development and validation of a stability-indicating UV spectroscopic method for Ilaprazole in bulk and pharmaceutical formulations ". Inventi Rapid. 2013.
10.Mansuri JC, Hariyani H, Kanani B and Faldu S. "Development and validation of spectrophotometric method for the determination of Ilaprazole in bulk drug and its pharmaceutical dosage form ".
11.Padhiyar P, Padaliya H, B Sanjaysingh, Patel H and Pancholi S.S. "Development and validation of UV spectrophotometric method for estimation of Ilaprazole in bulk and pharmaceutical dosage form ". Inventi Rapid. 2013.
12.B.satheesh, D.sravanan and K.gundu. "Simultaneous determination of Ilaprazole and its related compound in pharmaceutical dosage form by UPLC". J. Liq. Chromatogr. Relat. Technol. 2013, 20, 2968-2981.
13.Zhou TZ, Zhang W, Ou-Yang DS, Chen Y, Guo D, Liu YZ, Fan L and Deng HW. "An improved LC-MS/MS method for quantitative determination of Ilaprazole and its metabolites in human plasma and its application to a pharmacokinetic study". Actapharmaconsin.,30, 2009, 1330-1336.
14.Shah R, Talavia B, Patel P, Tailor P and ShahS, AICTE Sponsored seminar on Quality Assurance Issues in Drug Discovery & Formulation Development. 2013,
15.Lakshamanaprabu As, shirwaikar A, C DK, Joseph A and kumar R. "Simultaneous determination of Esomeprazole and Domperidone by UV spectrophotometric method". Indian J. pharm sci., 70, 2008, 128-131.
16.Kalure SU, Kulkarni R and SomwanshiGP."Spectrometric simultaneous determination of Domperidone and Pantoprazole". Der. Pharm. Chemica. 2012, 4, 1517-1521.
17.Jadhav K, Dhamecha DL, Ghadlinge SV, Asnani GP and PatilMB,“Simultaneous determination of Lafutidine and Domperidone by ultraviolet spectroscopy". ISRN Anal. Chem.,2012, 2012, 1-4.
18.Kumar YD, V Ravinder and M Rajesh. "Simultaneous eastimation of Omeprazole magnesium and Domperidone tablets by ultraviolet spectroscopy". IJPS, 2, 2011, 207-210.
19.Kalra K, Naik S, Jarmal G and Mishra Y. "Spectrophotometric method for simultaneous estimation of Paracetamol and Domperidone in tablet formulation". Asian J. Research Chem., 2009, 2, 112-114.
20.Patel AH, Patel JK and Patel KN. "Development and validation of derivative spectrophotometric method for simultaneous estimation of Domperidone and Rabeprazole sodium in bulk and dosage forms". Int.J.Ph.Sci., 2010, 464-469.
21.Rana S , Pandya J, Solanki S and Patel M. "Development and Validation of Spectrophotometric method for Simultaneous estimation of Lafutidine and Domperidone in combined dosage form by area under curve method". Ijddr, 2012, 4, 257-262.
22.Tarkase KN, Tarkase MK, Dokhe MD and Wagh VS. "Development and validation of spectrophotometric method for simultaneous estimation of Cinnarizine and Domperidone maleate in pure and tablet dosage form".Ijpsr, 2012, 3, 2700-2704.
23.ICH Q1 A (R2): Stability Testing of New Drugs and Products. International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use, ICH Harmonised Tripartite Guideline, 2003.
24.ICH Q2 (R1): Validation of Analytical Procedures: Text and Methodology. International Conference on Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human use, ICH Harmonised Tripartite Guideline, 2005.

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