ASSAY METHOD VALIDATION PROTOCOL OF SILDENAFIL CITRATE TABLET IP

ASSAY METHOD VALIDATION PROTOCOL OF Sildenafil Citrate Tablets IP

Superseded Protocol No.	Nil
Effective Date

Sr. No.	Subject
	Protocol Approval
	Objective
	Scope
	Responsibility of validation team
	Product profile
	Methodology
	Revision History

1. Protocol Approval:

Prepared By:

Functional Area	Name	Signature	Date
Officer / Executive Quality Control

Reviewed By:

Functional Area	Name	Signature	Date
Manager QC
Manager QA

Approved By:

Functional Area	Name	Signature	Date
Head QA

Authorized By:

Functional Area	Name	Signature	Date
Head Quality

• Objective:

The objective of this validation is to validate the assay method of Sildenafil Citrate Tablets IPby considering the following parameters:

• Precision
• System Precision
• Method Precision
• Intermediate Precision (Ruggedness)
• Specificity
• Forced Degradation
• Linearity
• Accuracy
• Range
• Stability in Analytical Solutions
• Filter Paper Selection Study
• Robustness  

Following parameters will be seen during system suitability:

Sr. No.	Parameters	Limit
1.	The Tailing Factor	Not more than 2.0
2.	% RSD for peck area of Sildenafil Citrate Tablets IP from 6 replicate injections of standard solution.	Not more than 2.0

When the above parameters meets the method, the actual experiment shall be started.

• Scope :

This protocol is based on SOP and applicable for the validation of assay method of Sildenafil in Sildenafil Citrate tablet.

• Responsibility of Validation Team:

Departments	Responsibilities
QC	Preparation & Review of Protocol.
Analysis of samples and recording of data.
Compilation and checking of data
Preparation of Summary Report.
To impart training of protocol to concerned department/persons.
QA	Review and approval of protocol.
Co-ordination with QC to carryout Validation.
Review of data and summary report.
Head Quality	Authorization of protocol.

• Product Profile:

Category (% API Content)
Reason for Validation
Active Ingredient	Sildenafil Citrate
Strength	Each Film Coated tablet contains: Sildenafil Citrate Eq. to Sildenafil 50 mg
Methodology	Pharmacopeial / Non-Pharmacopoeial / Provided by customer
Method Reference	IP
Specification Limits (Shelf Life)	90 % -110 % 24 Months

• Methodology:
  • Precision:
    • System Precision:

Test Procedure:

Prepare standard solution as per test method and inject for 6 times.

Documentation:

Record the retention time, peak area, and peak height on the datasheet. Calculate the mean, standard deviation, and RSD.

Acceptance criteria

% RSD shall be not more than 2.

• Method Precision:

Test Procedure:

Prepare six samples and analyze as per finished product testing procedure.

Documentation:

Record the % Assay of Sildenafil in Sildenafil Citrate Tablet on the datasheet. Calculate the mean, standard deviation, and % RSD.

Acceptance criteria

% RSD shall be not more than 2.

• Intermediate Precision (Ruggedness):

Test Procedure:

Prepare six samples and analyse as per finished product testing procedure by different analyst by using different columns, different systems on different day. Evaluate the system suitability criteria.

Documentation:

Calculate the mean, standard deviation, and % RSD for the operators and instruments and record on data sheet.

Acceptance criteria

% RSD shall be not more than 2.

• Specificity:

Test Procedure:

Prepare and inject blank (diluents), standard, sample & placebo.

Documentation:

Print chromatograms.

Acceptance criteria

The excipient compounds must not interfere with the analysis of the targeted analyte. No peak shall be eluted at the retention time of Sildenafil Citrate in blank and placebo.

• Forced Degradation:

Test Procedure:

Stress the sample at the following conditions and evaluate the peak purity.

• Degradation by Hydrochloric Acid
• Degradation by Sodium Hydroxide
• Degradation by hydrogen peroxide
• Degradation by thermal
• Degradation by water hydrolysis

Documentation:

Print chromatograms.

Acceptance criteria

Peak purity shall be passed. The difference between treated and untreated sample shall be not more than 10 %.

• Linearity:

Test Procedure:

Prepare the standard solutions at six concentrations, typically 25, 50, 75, 100, 150, and 200% of target concentration following finished product testing procedure. Three individually prepared replicates at each concentration will be analyzed.

Documentation:

Record results on a datasheet. Calculate the mean, standard deviation, and Relative Standard Deviation (RSD) for each concentration. Plot concentration (x-axis) versus mean response (y-axis) for each concentration. Calculate the correlation coefficient (r). Record these calculations on the datasheet.

Acceptance criteria

The correlation coefficient (r) shall be not less than 0.99.

• Accuracy:

Test Procedure:

Prepare sample solution by spiking the Sildenafil Citrate (Drug) with placebo at about 50%, 100% and 150% of test concentration and analyze as per finished product testing procedure.

Documentation:

For each sample, record the theoretical value, assay value, and percent recovery on test data sheet. Calculate the mean, standard deviation, RSD, and percent recovery for all samples. Record results on the datasheet.

Acceptance criteria

The mean recovery at each level shall be within ……….. to ………….% of the theoretical value and the % RSD not more than 2.

• Range:

Test Procedure:

The data obtained during the linearity and accuracy studies will be used to assess the range of the method. The precision data used for this assessment is the precision of the three replicate samples analyzed at each level in the accuracy studies.

Documentation:

Record the range on the datasheet.

Acceptance criteria

The acceptable range shall be defined as the concentration interval over which linearity and accuracy are obtained as per the above criteria.

• Stability in Analytical Solutions;

Test Procedure:

Prepare standard and one sample solution as per finished product testing procedure and inject into HPLC at initial and different time interval up to 48 hours and determine the % assay.

Documentation:

Calculate the % assay.

Acceptance criteria

The difference in the results shall be not more than 2 %.

• Filter Paper Selection Study :

Test Procedure:

Prepare sample solution in triplicate as per the finished product testing procedure and filter test with 0.45m PVDF filter and 0.45m Nylon filter and centrifuge a portion of filter.

Documentation:

Calculate the % assay of centrifuged and filtered samples.

Acceptance criteria

The difference in the results shall be not more than 2%.

• Robustness:

Test Procedure:

Prepare the sample solution in triplicate and analyze under the following conditions as per finished product testing procedure.

• By changing the flow rate by ±10%
• By changing the temperature by ±5°C
• By changing the organic content by ±10%
• By changing the pH of ………………. In mobile phase by ±0.2 units

System suitability shall be done in each condition.

 Documentation:

Calculate the % assay. Calculate the % RSD for % assay for individual above experiment with method precision results. 

Acceptance criteria

Overall % RSD shall be not more than 2.

• Revision History:

Revision No.	Details of changes	Reason