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

Revision No.: 00
REF. SOP NO.: 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):

Observed weight in gm.

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

(W2 – W1) / 10
Pump-B
Weight of beaker + water
after 10 min. (W2)
Weight of beaker (W1) 0.99 to 1.01

(W2 – W1) / 10

Weight per ml of water = ________ gm.

Calibrated By: Checked By:

Date Date :

ANNEXURE – 2

FORMAT FOR CALIBRATION RECORD OF HPLC

Revision No.: 00
REF. SOP NO.: 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.

Height From Base Line To Actual Value Set

Tine Range Value Obtained
B Concentration Peak. 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

Revision No.: 00
REF. SOP NO.: 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)  250 ml.  5ml  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 l.
Inject six replicate injection of standard solution and calculate RSD for area of main
peak.
Injection no. Area % of RSD Limit
1
2
3 Not More Than 1.0
4 %
5
6

2.Check for Linearity :

Correlation coefficient
Sr. Injection
Area Mean Observed
No. volume Limit
results
i)
1 10 l.
ii)
i)
2 20 l.
ii)
i) Not less than
3 30 l.
ii) 0.99
i)
4 40 l.
ii)
i)
5 50 l.
ii)
Remarks: Satisfactory / Not satisfactory.
Calibrated By: Checked By:
Date Date :

ANNEXURE – 4

FORMAT FOR CALIBRATION RECORD OF HPLC

Revision No.: 00
REF. SOP NO.: 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.) 250 ml.  5ml  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 l.

Sr.No. Wavelength (nm) Area Mean

1)
1 252
2)
1)
2 254
2)
1)
3 256
2)
1)
4 258
2)
1)
5 260
2)
1)
6 262
2)
1)
7 264
2)
Largest peak response is observed at __________ nm. (Limit: 258 nm ± 2 nm)
Remarks: Satisfactory / Not Satisfactory.

Calibrated By: Checked By:

Date Date :