FORMAT FOR CALIBRATION RECORD OF HPLC

Annexure – 1    :  Format For Calibration Record Of Pump (Flow Rate)

Annexure – 2    :  Format For Calibration Record Of Gradient Proportionate Valve

Annexure – 3    :  Format For Calibration Record Of Injector

Annexure – 4    :  Format For Calibration Record Of Detector

ANNEXURE – 1

FORMAT FOR CALIBRATION RECORD OF HPLC

REF. SOP NO.:	Revision No.:  00	Page No.: 1 of 1
	Effective from:

Calibration Record     :           High Performance Liquid Chromatograph

Instrument No.           :

Make And Model       :

Calibration Date         :

Calibration Due On    :

Calibration Of Pump (Flow Rate):

Flow rate adjusted to 1.0 ml / min.	Observed weight in gm.				Limit in ml / min.
	1	2	Mean wt. / min	Mean Vol. / min
Pump-A
Weight of beaker + water after 10 min. (W2)					0.99 to 1.01
Weight of beaker (W1)
(W2 – W1) / 10
Pump-B
Weight of beaker + water after 10 min. (W2)					0.99 to 1.01
Weight of beaker (W1)
(W2 – W1) / 10

Weight per ml of water =        ________ gm.

Calibrated By:                                                            Checked By:  

Date                                                                            Date : 

ANNEXURE – 2

FORMAT FOR CALIBRATION RECORD OF HPLC

REF. SOP NO.:	Revision No.:  00	Page No.: 1 of 1
	Effective from:

Calibration Record     :           High Performance Liquid Chromatograph

Instrument No.           :

Make And Model       :

Calibration Date         :

Calibration Due On    :

Calibration Of Gradient Proportionate Valve

Parameters :

Mobile Phase              : Reservoir  A 100% Water with HPLC

Reservoir B 0.3% Acetone HPLC 

Wavelength                 : 254 nm.

Flow rate                     : 2 ml / min.

Tine Range	Height From Base Line To B Concentration Peak.	Value Obtained	Actual Value Set In The Program
0.01 To 10.0		(Lt: 9.9 To 10.1)
10.01 To 20.0		(Lt: 49.5 To 50.5)
20.01 To 30.0		(Lt: 89.1 To 90.9)

Value  :      Height of B Concentration

                             --------------------------------------  

                  Height of a 100 % Concentration              

Remarks:    Satisfactory  / Not Satisfactory

Calibrated By:                                                            Checked By:  

Date                                                                            Date :

ANNEXURE – 3

FORMAT FOR CALIBRATION RECORD OF HPLC

REF. SOP NO.:	Revision No.:  00	Page No.: 1 of 1
	Effective from:

Calibration Record     :           High Performance Liquid Chromatograph

Instrument No.           :

Make And Model       :

Calibration Date         :

Calibration Due On    :

Calibration Of Injector : 1. Check for Precision:

Standard Preparation:

Take ______ mg of Uracil (about 50 mg) à 250 ml. à 5ml à 50ml with Methanol.

Parameters:

Mobile Phase : Methanol HPLC grade.       Column  :  ODS (150 mm x  4.6 mm. 5 micron)

Wavelength  :  254 nm.      Flow rate :  1 ml / min.    Injection volume : 20 ml.

Inject six replicate injection of standard solution and calculate RSD for area of main peak.

Injection no.	Area	% of RSD	Limit
1			Not More Than 1.0 %
2
3
4
5
6

2.Check for Linearity :

Sr. No.	Injection volume	Area	Mean	Correlation coefficient
				Observed results	Limit
1	10 ml.	i)  ii)			Not less than 0.99
2	20 ml.	i) ii)
3	30 ml.	i) ii)
4	40 ml.	i) ii)
5	50 ml.	i) ii)

Remarks: Satisfactory / Not satisfactory.

Calibrated By:                                                            Checked By:  

Date                                                                            Date :

ANNEXURE – 4

FORMAT FOR CALIBRATION RECORD OF HPLC

REF. SOP NO.:	Revision No.:  00	Page No.: 1 of 1
	Effective from:

Calibration Record     :           High Performance Liquid Chromatograph

Instrument Sr. No.                  :

Make And Model       :

Calibration Date         :

Calibration Due On    :

Calibration Of Detector

Standard Preparation:

Take _____ mg of Uracil ( about 50 mg.) à250 ml. à 5ml à 50ml with Methanol.

Parameters:

Mobile Phase:  Methanol HPLC grade       Column:  ODS (150 mm x 4.6 mm. 5 micron)

Wavelength:  252- 264 nm with 2 nm increment.      Flow Rate:  1 ml / min.    Injection Volume: 20 ml.

Sr.No.	Wavelength (nm)	Area	Mean
1	252	1) 2)
2	254	1) 2)
3	256	1) 2)
4	258	1) 2)
5	260	1) 2)
6	262	1) 2)
7	264	1) 2)

Largest peak response is observed at __________ nm. (Limit: 258 nm  ± 2 nm)

Remarks: Satisfactory / Not Satisfactory.

                                 

Calibrated By:                                                            Checked By:  

Date                                                                            Date :