Abstract

An easy-to-use, sensitive and accurate RP-HPLC technology has been developed and validated to detect Vonoprazan and Domperidone simultaneously in pharmaceutical dosage forms. The acetonitrile-based mobile phase buffer is supplied at a flow rate of 1.0 ml/min at a ratio of 65:35, with a detection wavelength of 230 am. The retention durations for vonoprazan and domperidone were 7.185 and 11.365 minutes, respectively. The method was developed and approved in accordance with ICH guidelines.

Keywords

Vonoprazan, Domperidone, RP-HPLC, Validation

Introduction

From the fields of development and validation, quality control, and quality assurance, the ongoing and interdependent tasks include the creation of analytical methods and validation. Analytical methods are essential for risk management and equivalency assessments. It supports the stability of outcomes and the development of acceptability criteria. The analytical procedure's suitability for the intended use should be demonstrated through validation. An essential tool for characterizing and validating a method is experiment design. To optimize the analytical method, it should be easy for analytical professionals to employ. Significant gains in accuracy and a decrease in errors can be obtained by well-developed analytical methods and their validation. Avoiding time-consuming and expensive exercises can be effective.[1] Vonoprazan fumarate 1-[5-(2-fluorophenyl)-1- ( pyridine -3 -yl sulfonyl ) – 1 H-pyrrol- 3 – yl ]- N - methyl methanamine fumarate , a chemical derivative of pyrroles, The potassium-competitive acid blocker Vonoprazan prevents acid secretion by competitively preventing potassium from being available to hydrogen-potassium ATPase. When fasting or fed, Vonoprazan is rapidly absorbed and acid stable, taking 1.5–2.0 hours to reach its Cmax. It slowly separates from its target, with a half-life of roughly 7.7 hours[3] Due to its high pKa (>9), it is more likely to accumulate in the canalicular region of parietal cells, where it inhibits resting and active proton pumps in a competitive manner[2] There is interindividual variation in the effects of CYP2C19, age, sex, and dose[2] It is not advised to change the dosage in cases of hepatic or renal illne

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        Structure of Vonoprazan

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Peripheral dopamine2-receptor antagonist domperidone has been demonstrated to have some effects on the motor function of the oesophagus in addition to regulating the motility of gastric and small intestine smooth muscle. Because it blocks dopamine receptors in the chemoreceptor trigger zone, it also possesses antiemetic properties. In patients with symptoms of diabetic gastropathy, domperidone relieved symptoms (anorexia, nausea, vomiting, abdominal pain, early satiety, bloating, and distension) more effectively than placebo in controlled clinical trials; Additionally, dopamine-containing medications avoided the gastrointestinal and emetic side effects of antiparkinsonian therapies, prevented nausea and vomiting linked to emetogenic chemotherapy, and temporarily relieved symptoms in patients suffering from dyspepsia or gastroesophageal reflux.  Domperidone barely penetrates the blood-brain barrier, therefore reports of negative effects on the central nervous system, like dystonic responses, are rare.

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    Structure of Domperidone

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MATERIALS AND METHODS

RP-HPLC method

Materials and reagents

Vonoprazan and Domperidone from Sai Mirra Pharma company, Chennai, Tamilnadu, India were received as a gift sample. Acetonitrile of HPLC grade from Qualigens fine chemicals, Mumbai, India. All the reagents and chemicals used were of analytical and HPLC grade. Water (HPLC grade) was obtained from Milli Q RO system.

Instruments and Chromatographic Conditions

Shimadzu HPLC system was used for method development and validation. Data acquisition was performed on Data ace HPLC software. The separation were achieved on C8 (250 × 4.6 mm, 5?m) column at ambient temperature and the eluent was monitored at 230 nm with flow rate of 1.0 mL/min using UV detector. The mobile phase mixture of 35 parts Acetonitrile & 65 parts Water was filtered through nylon 0.22 ?m membrane filters and was degassed before use (30 min). A volume of 20 ?L of each sample was injected into column.

Preparation of Buffer:

4.14 g of Sodium dihydrogen orthophosphate in 1000 mL of water adjust pH 6.5 with Sodium hydroxide or Ortho phosphoric acid.

Diluent:

Water: Acetonitrile (65:35)

Preparation of standard solution

Weigh accurately 67 mg of  Vonoprazan fumarate WRS and 25 mg Domperidone WRS transfer to a 50 ml volumetric flask. Dilute 5 mL into a 50 ml with diluent. Dissolve and make up the volume with mobile phase

Sample preparation :

Transfer 210 mg of sample to a 100 ml volumetric flask, add 70 ml of diluents and sonicate for 25 minutes with intermittent shaking. Allow to cool and dilute to volume. Filter through 0.45 µm membrane filter. Dilute 5 ml into a 10 ml with diluent.

Procedure:

Separately inject 20 µL blank, 6 replicates of the standard solution and check the system suitability. If the system suitability passes the acceptance criteria, proceed with the injection of sample solutions.

System suitability:

Relative standard deviation: NMT 2.0% for the 6 areas of Vonoprazan and Domperidone peaks from 6 replicate standard injections.

Column efficiency:

Not less than 2500 theoretical plates for both Vonoprazan and Domperidone

Resolution: Not less than 5.0 between Vonoprazan and Domperidone peaks. Calculate the content of Vonoprazan per tablet using the formula :

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Where, P =  Purity of  Vonoprazan fumarate WRS

Calculate the content of Domperidone  in % of label claim using the     

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P= Purity of Domperidone WRS              

METHOD VALIDATION

Procedure:

Separately inject the standard solution in 6 replicates and sample solution and record the chromatograms. The order of elution will be as follows:

VONOPRAZAN

DOMPERIDONE

Linearity:

The calibration curve was plotted between concentration and peak area. Vonoprazan and Domperidone were linear in the concentration range of 50-150mcg/ ml and 25-75 mcg/ ml respectively. All the solution were filtered through 0.22 µm membrane filter Aliquots (20 ?l) of each solution were injected under the operating chromatographic condition described above [Number of replicates (n = 6)]. A calibration graph was plotted between the mean peak area Vs respective concentration and regression equation was derived.

System suitability

The developed procedure was verified in compliance with ICH guidelines [7] Measurements were made of the system suitability characteristics in order to assess the system performance. The precision of the system was assessed using six identical injections of the same setup. Asymmetry and the number of theoretical plates were measured [8,9]. By calculating the peak retention time, peak area, theoretical plates, tailing or asymmetry factor for Vonoprazan and Domperidone. the system's appropriateness was ensured.

Relative standard deviation:

NMT 2.0% for the 6 areas of  Vonoprazan and Domperidone peaks from 6 replicate standard injections.

Column efficiency:

Not less than 2500 theoretical plates for both Vonoprazan and Domperidone were used for determining the system suitability.

Resolution:

Not less than 5.0 between Vonoprazan and Domperidone peaks

Precision

The reproducibility of an analytical procedure under typical operating conditions is known as precision. Repeatability (intra-day) and intermediate precision (inter-day) were used to determine the assay's precision; the results were expressed as a percentage R.S.D. for a statistically significant number of replicate readings. By comparing the assays on three separate days, the intermediate precision was investigated, and the results were recorded as standard deviation and % R.S.D. The % RSD for the six Assay determinations is NMT 2.0% were used for determining the precision

Accuracy

Recovery studies verified the method's accuracy. A known amount of standard Vonoprazan and Domperidone were added to the previously examined formulation, and the recovery % was computed. The recovery at various levels is between 98.0% and 102.0% ii. The RSD for Recovery of triplicate samples at various levels is not more than 2.0% were used for determining the Accuracy

Specificity

The ability to accurately and specifically measure the target analyte in the presence of other substances that would be expected to be present in the sample matrix is known as specificity. It measures the level of interference from other active ingredients and excipients, among other things, to make sure that a peak reaction is exclusively caused by one ingredient and that there is no co-elution. Specificity parameters were assessed using standard, blank, and placebo solutions to see whether Vonoprazan and Domperidone retention times were impacted in any way. There should be no interference of the placebo which determines the specificity.

RESULTS AND DISCUSSION

Linearity:

For linearity, Construction of the calibration curves for the drugs Vonoprazan and Domperidone are assessed by the combined standard solutions in range of 50-150mcg/ml and 25-75mcg/ml respectively. The responses were linear throughout the range for all two analytes {r?2;=0.999,r?2;=0.997,respectively}

Correlation Coefficient square – NLT

0.995 y-intercept : Not more than ± 2.0%

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Calibration curve of Domperidone

       
           
       
    

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System suitability:

In the present HPLC method, Acetonitrile and distilled water (65:35 v/v) were used as mobile phase. In a preliminary separate analysis study of Vonoprazan and Domperidone  C8 column was successfully used; therefore, C8 was used simultaneous determination. The described chromatographic conditions resulted in Vonoprazan and Domperidone  retention at about 7.167 and 11.256  minutes,

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Precision:

The reproducibility of an analytical procedure under typical operating conditions is known as precision. Repeatability (intra-day) and intermediate precision (inter-day) were used to determine the assay's precision; the results were expressed as a percentage R.S.D. for a statistically significant number of replicate readings. By comparing the assays on three separate days, the intermediate precision was investigated, and the results were recorded as standard deviation and percent R.S.D.

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Accuracy:

According to FDA, percentage recovery method was used to determine Accuracy. The recovery at various levels is between 98.0% and 102.0% ii. The RSD for Recovery of triplicate samples at various levels is not more than 2.0% were used for determining the Accuracy

For Vonoprazan:

20 mg  drug solutions was taken in three different flask label A1,A2,A3.spiked 50%,100%,150% of standard solution in it and diluted .the area of each solution at 230 nm. the amount of vonoprazan was calculated at each  level and percentage recoveries were calculated.

For Domperidone:

10 mg drug solution was taken in three different flask label A1,A2,A3. Spiked 50%,100%,150% of standard solution in it and diluted. The area of each solution  at 230 nm. The amount of domperidone was calculated at each level and percentage recoveries were calculated.

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Vonoprazan

       
           
       
    

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Data of Accuracy of Vonoprazan

       
           
       
    

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Data of Accuracy of Domperidone

Remarks: The recovery and RSD for recovery at each level meets the acceptance criteria

Specificity:

The specificity of the drug Vonoprazan and Domperidone were assessed using standard, blank, placebo solutions which impacts the retention time.

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Remarks:

There is no interference of the placebo. Hence the method is specific

CONCLUSION

In accordance with ICH guidelines, the current RP-HPLC developed method for the simultaneous evaluation of Vonoprazan and Domperidone was verified. It was discovered that the developed RP-HPLC method was easy to use, fast, accurate, selective, and precise for the simultaneous quantification of all two drugs. Because the validated approach yielded no incorrect findings, it can be utilized for routine examination of two drugs.

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