amv report for paracetamol injection

Name of Product	PARACETAMOL INJECTION
Parameter	Analytical Method Validation for Paracetamol Injection by HPLC
Method followed	By HPLC
Supersede No.	Nil
Total No of pages	19
Effective Date

TABLE OF CONTENTS

Sr. No.	Title	Page
—	Product Description
—	Table of contents
1.0	Document Preparation and Protocol Approval
2.0	Revision Control History
3.0	Objective
4.0	Scope
5.0	Reason For Validation
6.0	Responsibility
7.0	Description of Method Belonging
8.0	Training Details
9.0	Details of Analytical Method
10.0	Chromatographic Conditation
11.0	Performance Parameter of Validation
12.0	Selection of Analytical Parameter Performs
13.0	Specificity (Selectivity)
14.0	Precision
15.0	Table for sample assay
16.0	Inter Mediate Precision
17.0	Accuracy (Recovery)
18.0	Sample Sequence of Accuracy
19.0	Linearity
20.0	Robustness
21.0	Normal Condition
22.0	Sample Sequence of Robustness
23.0	Acceptance criteria
24.0	References

1.0 DOCUMENT PREPARATION AND PROTOCOL APPROVAL

Preparation and Approval of this Analytical Method Validation document will be joint responsibility of the following functional area. Any modification in this document shall be documented and approved.

1.1 Prepared By:

1.2 Reviewed By:

1.3 Approved By:

2.0 Revision Control History:

3.0 OBJECTIVE:
To perform Analytical method Validation of Paracetamol Injection and provide documentary evidence that the method used for Analytical method Validation of said product full fill the regulatory & compendia requirement. This protocol presents and discusses the result obtained in the Verification study for the parameter namely Specificity, System Suitability, Precision (Repeatability), Accuracy, Linearity, Robustness and the test method.

4.0 SCOPE:
This document is applicable for the Analytical method Validation of HPLC Method which is applying for the Determination of Analytical validation for Paracetamol Injection, to be manufactured at
Fast Pharma Pvt. Ltd.

5.0 REASON FOR VALIDATION:
Regulatory Requirement

6.0 RESPONSIBILITY:
The following functions are involved in the Analytical method Validation performing & shall carry out the following responsibilities.

6.1 Quality Control:
6.1.1 Officer & Executive Quality Control: To perform the analysis in the Verification and evaluate the data as per protocol.
6.1.2 Sr. Officer: To prepare the protocol and review all data evaluated as per protocol.
6.1.3 Manager Quality Control: To provide technical support, review the protocol and ensure that all parameter performed as per protocol.

6.2 Regulatory:
6.2.1 To provide technical support, review and approved the protocol and ensure that all parameter performed as per protocol.

7.0 DESCRIPTION OF METHOD BELONGING:
7.1 Product Detail:
7.1.1 Name of Product: PARACETAMOL INJECTION
7.1.2 Generic Name: PARACETAMOL INJECTION
7.2 Method Details:
Methodology: High performance liquid chromatography

8.0 TRAINING DETAILS:
Training shall be imparted to all concern persons involved in Analytical method validation activity w.r.t. cGMP & concerned analytical technique required for analysis & details will be attached in analytical method verification report’ activity, as per below mentioned format

Purpose	To train all personnel involved in the execution of this protocol
Topic	· Purpose · Procedure · Method details · Acceptance criteria · Documentation etc.
S. No.	Name of Participants	Area of Operation	Sign/Date

Name of the Trainer: _______________

(Sign, Date & Time):

9.0 DETAILS OF ANALYTICAL METHOD:

9.1 Type of Analysis: Liquid chromatography

9.2 Equipment/Instrument required during validation

Sr. No.	Name of Equipment	Equipment No.	Calibration Status		Make/Model
Sr. No.	Name of Equipment	Equipment No.	Calibration on	Calibration Due	Make/Model
1.	Analytical Balance
1.	Analytical Balance
2.	HPLC with UV Detector
2.	HPLC with UV Detector
3.	Column
4.	pH Meter
5.	Sonicator
6.	Filtration assembly
7.	Filters

9.3 Reagents and Chemicals

Sr. No.	Name of Reagent	Grade	Make	Lot No.
1.	Orthophosphoric Acid

Diluents: Buffer and Acetonitrile (30:70)
Standard solution:
Dissolve 150 mg of Paracetamol RS in the mobile phase and dilute to 100 ml with the mobile phase. Dilute 5 ml of this solution to 50 ml with the mobile phase.
Test solution:
Dissolve (1ml) 150 mg of Paracetamol RS in the mobile phase and dilute to 100 ml with the mobile phase. Dilute 5 ml of this solution to 50 ml with the mobile phase.

9.4 Chromatographic Condition:
Column Dimension: a stainless steel column 25 cm × 4.6 mm, packed with octadecylsilane bonded to porous silica (5µm),
Buffer: Dissolve 2.5 ml Orthophosphoric acid diluted to 1000 ml with water.
Mobile phase: Buffer and Acetonitrile (30:70).
Flow rate: 1.0 ml per minute.
Wave length: 272 nm
Injection volume: 20 µl.
System suitability:
Samples: Inject the Standard solution and the test solution
Suitability requirements:
Relative standard deviation: ……………….not more than 2% for Paracetamol standard solution.
Tailing Factor: …………….. Not more than 2.0, Standard solution
Analysis Samples: Standard solution and Sample solution

9.5 Calculation:

10.0 Performance Parameter of Validation:

Sr. No.	Performance Parameter
1.	Specificity, System Suitability, méthode Precision (Repeatability) System precision and Inter mediate precision, Accuracy, Linearity and Robustness.

11.0 SELECTION OF ANALYTICAL PARAMETER PERFORMS:

To the validation of the HPLC method used for the determination of Assay of Paracetamol Injection the following parameters should be performing.

11.1 Specificity (Selectivity):

Purpose: To establish Specificity of test method and to demonstrate its ability to assess unequivocally the Analyte in the presence of components which may be expected to be present? It ensures that there is no interference from Excipients and /or degradation products and /or impurities or the Analyte with each other.
Study Design: Demonstrate the Specificity of the test method by Injecting Blank, Standard solution & test solution.
Chromatographic Condition: …………………………………………………As per methodology.
Blank: …………………………Diluent
Diluents: ………………………………………Buffer and Acetonitrile (30:70)
Standard solution: Dissolve _{————-} 150 mg of Paracetamol RS in the mobile phase and dilute to 100 ml with the mobile phase. Dilute 5 ml of this solution to 50 ml with the mobile phase.
Test solution: Dissolve _{————-} (1ml) 150 mg of Paracetamol Injection in the mobile phase and dilute to 100 ml with the mobile phase. Dilute 5 ml of this solution to 50 ml with the mobile phase
The Observations are tabulated below

Sr. No.	Type of Solution	Retention Time	Area
1.	Blank
2.	Standard solution
3.	Test solution

Data Evaluation: Each chromatogram shall be processed & peak areas shall be calculated & reported.
Acceptance Criteria: RSD of five replicate injections should not be more than 2.0 %.
System Suitability:
- Tailing Factor: Not more than 2.0 for the Paracetamol peak, Standard solution.
- RSD: NMT 2.0%, Standard solution.
Conclusion: …………………………………………………………………………………………..

…………………………………………………………………………………………..

11.2 Precision:

The Precision of an analytical procedure is the degree of agreement among individual test results when the procedure is applied repeatedly to multiple samplings of a homogeneous sample. The precision of an analytical procedure is usually expressed as the standard deviation (SD) or relative standard deviation (RSD) (Coefficient of variation) of a series of measurements. Precision may be a measure of either the degree of reproducibility or of repeatability of the analytical procedure under normal operating conditions.

11.2.1 SYSTEM PRECISION:

Diluent, standard solution and chromatographic condition: Proceed as directed under point no. 9.0.
Procedure: five replicate injections of standard preparation shall be chromatogram as per analytical method.
Calculation: Calculate the relative standard deviation for replicate injections is not more than 2.0%.
Conclusion: ………………………………………………………………………………………..

11.2.2 METHOD PRECISION:

A) Repeatability:

Purpose: The Purpose of method Precision is to evaluate & demonstrate the precision of Analytical method under the same operating conditions over a short interval of time.
Study Design: The assay is determined of 6 test solutions at 100% concentration representing a precision of method evaluated by % RSD of assay Results.
Chromatographic Condition: ………………………………………………As per methodology.
Diluent, standard solution, chromatographic condition and procedure: Proceed as directed under point no. 9.0.
Test Solution: Six assay preparations of the sample shall be prepared as per the Analytical method.

The Observations are tabulated below

Sr. No.	Standard No.	Area
01.	Standard Solution -1
02.	Standard Solution -2
03.	Standard Solution -3
04.	Standard Solution -4
05.	Standard Solution -5
	Average
	%RSD

Observation Table for Sample and Assay:

Data Evaluation: The Relative Standard Deviation (RSD) of Assay of six preparations shall be calculated and reported.
Acceptance criteria: RSD of Assay of six preparations should not be more than 2.0 %.

System suitability:
- Tailing Factor: _____________ not more than 2.0 for the Paracetamol peak, system suitability solution.
- RSD: _____________ not more than 2.0%, Standard solution.
Conclusion: ………………………………………………………………………………………..

11.2.3 INTER MEDIATE PRECISION

Purpose: The Purpose of method Precision is to evaluate & demonstrate the precision of Analytical method under the same operating conditions over a short interval of time.
Study Design: The assay is determined of 6 test solutions at 100% concentration representing a precision of method evaluated by % RSD of assay Results.
Chromatographic Condition: ………………………………………………As per methodology.
Diluents, standard solution, chromatographic condition and procedure: Proceed as directed under point no. 9.0.
Test Solution: Six assay preparations of the sample shall be prepared as per the Analytical method.

The Observations are tabulated below

Sr. No.	Standard No.	Area
01.	Standard Solution -1
02.	Standard Solution -2
03.	Standard Solution -3
04.	Standard Solution -4
05.	Standard Solution -5
	Average
	%RSD

12.0 Accuracy (Recovery) :

Purpose: The Purpose of Accuracy of test method is to evaluate & demonstrate the closeness of agreement between the value which is accepted either as a conventional true value or an accepted reference value and the value found.
Study Design: The accuracy is demonstrated by preparing recovery samples (i.e. spiking known qty. of API) at a level of 50 %, 100 % & 150 % of sample concentration at Triplicate in each level.

Chromatographic Condition: As per methodology.
Diluent, standard solution, chromatographic condition and procedure: Proceed as directed under point no. 9.0.
Test Solution: Paracetamol Injection at three different levels 50 %, 100 % and 150 % of test concentration in triplicate (i.e. total nine determinations) covering the specified range and analyzed as described under methodology.
Sample Sequence for Accuracy:

Sample Name	Number of Injection	Injection Volume	Run Time
Blank	1	20µl
Standard solution	5	20µl
Test solution	3 different level (duplicate in each level)	20µl
Standard solution Bkt.	1	20µl

The Observations are tabulated below :

Sr. No.	Standard No.	Area
01	Standard Solution -1
02	Standard Solution -2
03	Standard Solution -3
04	Standard Solution -4
05	Standard Solution -5
	Average
	%RSD

The observation are tabulated in table:

Recovery level	Set No.	Peak Average area	Recovery (%)	Ave. (%)	RSD (%)
50%
100 %
150 %

Data Evaluation: The Relative Standard Deviation (RSD) of Assay of three preparations shall be calculated and reported.
Acceptance criteria: RSD of Assay of three preparations should not be more than 2.0 %.
System suitability:
- Tailing Factor: ______________ not more than 2.0 for the Paracetamol peak, system suitability solution.
- RSD: _________________ not more than 2.0%, Standard solution.
Conclusion: …………………………………………………………………………………………..

13.0 Linearity:

Purpose: The Purpose of Linearity testing of analytical procedure is to demonstrate its ability (within a given range) to obtain test results which are directly proportional to the concentration (amount) of analyte in the sample.
Study Design: Demonstrate the Linearity of the test method Blank & Stock solution (given below). A series of standard solutions shall be prepared (minimum of five preparations) over a range of 50 % to 150 % of the standard concentration
Chromatographic Condition: As per methodology.
Diluent, standard solution, chromatographic condition and procedure: Proceed as directed under point no. 9.0.
Standard solution: Dissolve _{————-} 150 mg of Paracetamol RS in the mobile phase and dilute to 100 ml with the mobile phase. Dilute 5 ml of this solution to 50 ml with the mobile phase the mobile phase.
Test solution: Dissolve _{————-} (1ml) 150 mg of Paracetamol Injection in the mobile phase and dilute to 100 ml with the mobile phase. Dilute 5 ml of this solution to 50 ml with the

Mobile phase.

Sample Sequence for Linearity:

Linearity Level	Sample Name with Conc. In µg/ml	Stock Solution	Diluted to volume	Injection Volume	No. of Injection	Run Time
—	Blank	—	—	20µl	01
—	Standard Solution	—	—	20µl	05
50 %	Linearity -75 ppm	2.5 ml	50 ml	20µl	01
80 %	Linearity -130 ppm	4.3 ml	50 ml	20µl	01
100 %	Linearity 150 ppm	5.0 ml	50 ml	20µl	01
120 %	Linearity -195 ppm	6.5 ml	50 ml	20µl	01
150 %	Linearity -225ppm	7.5 ml ml	50 ml	20µl	01

Data Evaluation: Inject above solution into HPLC System & record the area of Analyte peak. Plot a graph of concentration (in X-Axis) Vs Analyte peak area (in Y-Axis) and the Correlation coefficient reported.
Acceptance criteria: The Correlation coefficient should be not less than 0.99
System suitability:
Tailing factor: ______________ not more than 2.0 for the Paracetamol peak, system suitability solution.
RSD: ______________ not more than 2.0%, Standard solution.
Conclusion: …………………………………………………………………………………………..

14.0 Robustness:

Purpose: To establish the Robustness of Test method & to demonstrate its reliability by alter minimum two changes in critical parameter by minor variation.
Study Design: Demonstrate the Robustness of the test method by Carrying out System Suitability under normal condition & each of altered condition mention below.
Chromatographic Condition: As per methodology.
Mobile phase: As per methodology.
Blank:

i) Normal Condition:

Standard solution: Dissolve _{————-} 150 mg of Paracetamol RS in the mobile phase and dilute to 100 ml with the mobile phase. Dilute 5 ml of this solution to 50 ml with the mobile phase the mobile phase.
Test solution: Dissolve _{————-} (1ml) 150 mg of Paracetamol Injection in the mobile phase and dilute to 100 ml with the mobile phase. Dilute 5 ml of this solution to 50 ml with the

mobile phase.

The Observations are tabulated below for flow rate 1.0 ml/min. :

Sr. No.	Standard No.	Area
01	Standard Solution -1
02	Standard Solution -2
03	Standard Solution -3
04	Standard Solution -4
05	Standard Solution -5
	Average
	%RSD

Observation Table for Sample and Assay

Sample No.	Average area	Assay %
Normal Condition

ii) Flow rate: ± 20 % of Absolute (1.0 ml per minute),

Minimum Change : -20 % (0.8 ml/ minutes)
Maximum Change : +20% (1.2 ml/ minutes)
Standard solution: Dissolve _{————-} 150 mg of Paracetamol RS in the mobile phase and dilute to 100 ml with the mobile phase. Dilute 5 ml of this solution to 50 ml with the mobile phase the mobile phase.
Test solution: Dissolve _{————-} (1ml) 150 mg of Paracetamol Injection in the mobile phase and dilute to 100 ml with the mobile phase. Dilute 5 ml of this solution to 50 ml with the

Mobile phase. .

Sample Sequence for Robustness:

Sample Name	Number of Injection	Injection Volume	Run Time
Blank	1	20µl
System suitability solution	1	20µl
Standard solution	5	20µl
Test solution	2	20µl
Standard BKT	1	20µl

The Observations are tabulated below for flow rate 0.8 ml/min. :

Sr. No.	Standard No.	Area
01	Standard Solution -1
02	Standard Solution -2
03	Standard Solution -3
04	Standard Solution -4
05	Standard Solution -5
	Average
	%RSD

The Observations are tabulated below for flow rate 1.2 ml/min.:

Sr. No.	Standard No.	Area
01	Standard Solution -1
02	Standard Solution -2
03	Standard Solution -3
04	Standard Solution -4
05	Standard Solution -5
	Average
	%RSD

Observation Table for Sample and Assay:

Note: Same Analysis sequence will be applicable for each of the change conditions.
Data Evaluation: The standard deviation & Relative Standard Deviation (RSD) of area of five replicate injections of the standard preparation shall be calculated and reported.
Acceptance criteria: The % relative difference between assay values obtained in robustness study should not be not more than 2.0 %
System suitability:
Tailing factor: ……………… not more than 2.0 for the Paracetamol peak, system suitability solution.
RSD: ……………… not more than 2.0%, Standard solution.

Conclusion: ………………………………………………………………………………………….

14.0 ACCEPTANCE CRITERIA:

Sr. No.	Title	Acceptance Criteria		Observations/Conclusions
1.	Specificity	There shall be no interference from the peaks due to blank at the retention time of main ingredient.
2.	Method Precision
2.1	Repeatability		RSD of assay of six preparations should not be more than 2.0 %.
2.2	System precision		RSD of assay of six preparations should not be more than 2.0 %. The relative difference from method precision should be not more than 2.0 %.
2.3	Inter Mediate Precision		RSD of assay of six preparations should not be more than 2.0 %. The relative difference from method precision should be not more than 2.0 %.
3.	Recovery		Average recovery at each respective level within 95.0% to 105.0% and RSD of Triplicate recovery results should be not more than 2.0%.
4.	Accuracy (Recovery)		RSD of assay of 3 preparations should not be more than 2.0 %.
5.	Linearity		The Correlation coefficient should be not less than 0.999.

6.	Robustness		RSD of assay different condition should not be more than 2.0 %.