SOP for Linearity Check of High Performance Liquid Chromatograph (HPLC) : Pharmaceutical Guidelines

SOP for Linearity Check of High Performance Liquid Chromatograph (HPLC)

Standard operating procedure to check performance of the high performance liquid chromatograph (HPLC) for accurate results.

1.0 Objective

To check the performance of the equipment for reliable and accurate results.

2.0 Scope

This procedure is applicable to check the performance of the high-performance liquid chromatography.

3.0 Responsibility

3.1 Doing: Technical Assistant/Executive
3.2 Checking: Executive /Manager

4.0 Accountability

Head of the Department

5.0 Procedure

5.1 Operate the instrument as per the respective standard operating procedure of HPLC.
5.4 Instrument Performance ( Linearity of Results)
Frequency: Two months.
5.4.1 Prepare the mobile phase consisting of 70% Methanol: 30 % water.
5.4.1 Stock solution preparation: Prepare a mixture of 0.2 % Benzene and 0.2 % Toluene in methanol.
5.4.2 Make dilution in the following manner.
5.4.1 Take 5 ml, 10 ml, 15 ml, 20 ml of stock solution respectively in 25 ml of volumetric solution and makeup with methanol.
5.4.3 Set flow rate at 1.0 ml/min.
5.4.4 Allow the system to be saturated with mobile phase for at least 15 minutes before injecting the different solutions.
5.4.5 Record the area and retention time of both benzene and toluene as per Annexure - I.
5.4.6 Calculate the Linearity for both Benzene and Toluene respectively.

6.0 ABBREVIATION

6.1 SOP - Standard Operating Procedure

Related: Relative Response Factor Calculation in HPLC

ANNEXURE-I
Q. C. Department
LINEARITY CHECK REPORT OF HIGH-PERFORMANCE LIQUID CHROMATOGRAPH
PERFORMANCE DATE

DATE OF LAST PERFORMANCE
DONE
NEXT DUE FOR
PERFORMANCE



INSTRUMENT DETAILS

 INSTRUMENT NAME

INSTRUMENT MAKE

INSTRUMENT
IDENTIFICATION NO.



OBSERVATION
1) Instrument performance ( Linearity of results)
Stock solution preparation :
Mixture of 0.2 % Benzene and 0.2 % Toluene in Methanol.
Solution 1) : 5.0 ml of Stock solution                     25 ml with methanol.
Solution 2) : 10.0 ml of Stock solution                   25 ml with methanol.
Solution 3) : 15.0 ml of Stock solution                   25 ml with methanol.
Solution 4) : 20.0 ml of Stock solution                   25 ml with methanol.
Solution 5) : Direct stock solution.
HPLC Parameter :
Column         : ODS (25 cm X 4.6 mm, 5 µ)
Mobile phase : Methanol : Water
                            70     :   30
Flow rate        :  1.0ml/min.
Wave length    :  At 254 nm
Solution
No.
Conc.
Inj.No.
Bin No.
Area of Benzene
Area of Toluene
1
0.04%
1





2



2
0.08%
1





2



3
0.12%
1





2



4
0.16%
1





2



5
0.20%
1





2



Linearity : (Limit NLT 99.95%)







Ankur Choudhary is India's first professional pharmaceutical blogger, author and founder of Pharmaceutical Guidelines, a widely-read pharmaceutical blog since 2008. Sign-up for the free email updates for your daily dose of pharmaceutical tips.
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