sop for for hplc column receipt checking id no and regeneration

1.0 OBJECTIVE
1.1 To describe a procedure for Receipt, Checking, Id No. and Regeneration of HPLC column.
2.0 SCOPE
2.1 This procedure is applicable for Receipt, Checking, Id No. and Regeneration of HPLC column of Quality Control Department
3.0 RESPONSIBILITY
3.1 Officer /Executive – QC
4.0 ACCOUNTABILITY

4.1 Head – QC
5.0 PROCEDURE

5.1 On receipt of new column check the column received as per indent and mention details of column in Format

5.2 Ensure that the instrument is clean. Condition the column using methanol at 0.5 ml/minute for 30 minutes.

5.3 Check the column with respect to certificate of analysis for Back Pressure.

5.4 Carry out the column performance (Theoretical Plates, Tailing & Resolution) using following chromatographic condition. Record the results in Format

5.5 Performance of Reversed Phase Column

Chromatographic conditions:

Mobile phase : Methanol : water (70:30) % v/v

Flow rate : 1.0ml/min

Wavelength : 254nm

Inj. Volume : 20µl

5.6 Test solution for C8 and C18: Mixture of Benzene and Toluene (0.1%) in mobile phase.

5.7 After checking, flush the column with methanol for 30 minutes at flow rate of 0.5 ml/min. in normal direction and store the column in methanol.

5.8 Test solution for Cyano and Phenyl: Mixture of Benzene (0.1%) and Naphthalene (4mg) in 100 ml Mobile phase.

5.9 Performance of Normal Phase Column

5.10 Chromatographic conditions :-

Mobile phase : IPA : Hexane (40 : 60) v/v

Flow rate: 1.0ml/min.

Wavelength : 254nm

Inj. Volume : 20µl

Test solution: Mixture of Toluene (0.5 µl/ml) and Diethyl phthalate (2µl/ml) in mobile phase.

5.11 After checking, flush the column with Hexane for 30 min. at flow rate of 1.0ml/min and store the column in Hexane.

5.12 Enter the details of column in Format

5.13 Column Regeneration for Reverse Phase Column.

5.14 Flush the column using warm water (55ºC) at flow rate of 0.5 ml/min. for 20 minutes. Maintain the column at (55ºC) in column oven during flushing.

5.15 Inject 100 µl of DMSO four times at interval of 2 minutes.

5.16 Flush it with methanol at flow rate of 0.5 ml/min in reverse direction for 60 minutes. After flushing with methanol,

flush the column with chloroform for 60 minutes at flow rate of 0.5 ml/min in reverse direction.

5.17 At the end, wash the column in methanol for 30 minutes at flow rate of 0.5 ml/min. In reverse direction.

5.18 After regeneration of column, perform the column performance test as per Step 5.4.1.

5.19 Store the column in methanol, after flushing with methanol for 30 minutes at flow rate of 0.5 ml/min in normal direction.

5.20 Maintain the column History card on use of column during analysis as per Format

5.21 Column Regeneration for Normal Phase Column

5.22 Flush the column with Hexane for 60 minutes at flow rate of 2.0 ml/min.

5.23 After flushing with Hexane, wash the column with Methylene chloride for 60 min at flow rate of 2.0 ml/min

5.24 After flushing with methylene chloride, wash the column with Isopropyl alcohol for 60Minutes at flow rate of 2.0 ml/min.

5.25 At the end, flush the column with methanol for 60 minutes at flow rate of 2.0 ml/min.

5.26 After regeneration of column, perform the column performance test as per Step 5.4.2

5.27 Store the column in Hexane, after flushing with Hexane for 20 minutes at flow rate of 2.0ml/min in normal direction.

5.28 If the system suitability fails then only column regeneration required.

5.29 Column identification Number shall be given as for Column numbering will be: DB/QC/C/XXX
Where DB means Devbhoomi Pharmaceuticals.
QC: For Quality Control.
C: For Column.
XXX: Serial number starting from 001.
5.30

Column suitability parameters during CPT:

5.30.1 Theoretical Plates NLT 70% with respect to manufacturer COA.

5.30.2 Tailing between 0.8 to 1.5.

5.30.3 Resolution between two peaks NLT 5.

6.0 REFERENCE

6.1 In House

7.0 FORMATS
HPLC column check & regeneration record

Column Received & Identification No.

Log Book for Column

Column Index

8.0 ABBREVIATIONS
8.1 QC : Quality Control
8.2 QA : Quality Assurance
8.3 QCA : Quality Control Analytical
8.4 SOP : Standard Operating Procedure
8.5 No.: Number

8.6 Dept.: Department

8.7 min: Minute

8.8 °C : Degree Celsius

8.9 IPA: Isopropyl Alcohol

8.10 µl: Micro liter

8.11 DMSO: Dimethylsulfoxide