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Document Title : HPLC CALIBRATION

Reference SOP : PROCEDURE FOR OPERATION&CALIBRATION OF HPLC

Reference SOP No. : PB/SOP/QCD/001/01 Format No. : F/PB/QCD/001/01

Annexure No. : III Page No. : 1 of 1

HPLC ID No: ------------------------------- Column ID No : -------------------------

Make : ------------------------------ Model : --------------------------

Date of calibration: ------------------------ Calibration due Date: ---------------------------

Frequency : Half Yearly

1. FLOW RATE:

Weight per ml of water

Wt of Empty Pycnometer g

Wt / ml

=g/ml

Wt of Filled Pycnometer g

Wt of Water g 25.0

Set flow

Wt. of

empty

Cylender

(W1)

Wt. of

Cylender +

Water after

10 minutes

(W2)

Flow rate

(ml/min)=

[(W2-W1)/10 xwt/ml]

Average

flow Rate

(ml/min)

Limit + 2.0%

Minimum

(ml/min)

Maximu

m

(ml/min)

1.0 ml/min

g g

0.98 1.02g g

g g

2.0 ml/min g g 1.96 2.04

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Document Title : HPLC CALIBRATION

Reference SOP : PROCEDURE FOR OPERATION&CALIBRATION OF HPLC

Reference SOP No. : PB/SOP/QCD/001/01 Format No. : F/PB/QCD/001/01

Annexure No. : III Page No. : 2 of 1

2. Linearity (Detector) :

Reagent Name : --------------------------------------- Reagent B.No: -----------------------------

Mobile phase :70:30Methanol&Water Balance ID.No:------------------------------

Column : ----------------------------------- Column ID .No:-----------------------------

Wave length : ---------------------------------- Flow Rate (1ml/Min) :---------------------

Sample Solution A: Weigh accurately about ____(100 mg) of Caffeine Anhydrous Pure in 100 ml of

Volumetric Flask, dissolve and make up to mark with Mobile Phase.

Sample Solution B: dilute 1 ml of this solution A to 100 ml with Mobile Phase (10 ppm).

Sample Solution C:dilute 2 ml of this solution B to 100 ml with Mobile Phase (20 ppm).

Sample Solution D: dilute 3 ml of this solution C to 100 ml with Mobile Phase (30 ppm).

Sample Solution E: D dilute 4 ml of this solution D to 100 ml with Mobile Phase (40 ppm).

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Document Title : HPLC CALIBRATION

Reference SOP : PROCEDURE FOR OPERATION&CALIBRATION OF HPLC

Reference SOP No. : PB/SOP/QCD/001/01 Format No. : F/PB/QCD/001/01

Annexure No. : III Page No. : 3 of 1

S.NoConcentrati

onArea Mean Area

Correlation

Coefficient (r)

Acceptance

Criteria

01 10 ppm

0.98 to

1.000

02 20 ppm

03 30ppm

04 40ppm

05 50ppm

Calibrated by: ------------------- Checked by: --------------------

Date : ----------------- Date : -------------------

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8/11/2019 HPLC Calibration Format

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Document Title : HPLC CALIBRATION

Reference SOP : PROCEDURE FOR OPERATION&CALIBRATION OF HPLC

Reference SOP No. : PB/SOP/QCD/001/01 Format No. : F/PB/QCD/001/01

Annexure No. : III Page No. : 5 of 1

4. Wavelenght Accuracy:

Sample Solution A : Weigh accurately about ____(100 mg) of Caffeine Anhydrous Pure in 100 ml of

Volumetric Flask, dissolve and make up to mark with Mobile Phase.

Sample Solution B: Further dilute 2 ml of this solution to 100 ml with Mobile Phase (20 g/ml).

S.No Wavelength (nm) Area Maxima at Acceptance Criteria

1 270

273nm

2 271

3 272

4 273

5 274

6 275

7 276

Instrument Calibration meets the requirement /doesn`t meets the requirements

Calibrated by: ------------------- Checked by: --------------------

Date : ----------------- Date : -------------------

8/11/2019 HPLC Calibration Format

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Document Title : HPLC CALIBRATION

Reference SOP : PROCEDURE FOR OPERATION&CALIBRATION OF HPLC

Reference SOP No. : PB/SOP/QCD/001/01 Format No. : F/PB/QCD/001/01

Annexure No. : III Page No. : 6 of 1