1.0 OBJECTIVE:
To
lay down the operating procedure of UV/VIS Spectrophotometer.
2.0 SCOPE:
This
shall be applicable to the UV/VIS
Spectrophotometer of Quality Control Instrument Laboratory.
3.0 RESPONSIBILITY:
3.1
Technical
Assistant/ Helper.
3.2
Q.C Analyst.
4.0 ACCOUNTABILITY:
4.1
Manager-Quality
Control.
5.0 MATERIAL AND
EQUIPMENT:
5.1
UV/VIS Spectrophotometer.
6.0 PROCEDURE:
6.1
Ensure that the instrument is properly
connected to the stabilized power supply.
6.2
Switch ON the main
power supply switch.
6.3
The yellowish
light flickers and the instrument start INITIALISATION as under.
6.3.1
Lsi initialize.
6.3.2
Rom check.
6.3.3
Ram check.
6.3.4
Filter origin.
6.3.5
Light source org.
6.3.6
λ org. (coarse).
6.3.7
W lamp energy.
6.3.8
λ org (fine).
6.3.9
D2 lamp
energy.
6.3.10
Check.
6.4
After initialization
of the instrument push MODE key. The MAIN MENU appears on the screen as under:
6.4.1
Photometric.
6.4.2
Spectrum.
6.4.3
Quantitation.
6.4.4
Kinetics.
6.4.5
Time scan.
6.4.6
Multiple
components.
6.4.7
Optional program
pack.
6.4.8
UTILITIES.
6.5
Select
option:
6.5.1
For further
operation on the instrument any option as listed in point no. 6.4 is chosen.
6.6
Determination
of absorbance:
6.6.1
Set the cursor at
photometry.
6.6.2
Press the “GO TO” on the keyboard.
6.6.3
“GO
TO WL” [190.0-1100.0] appears on the screen.
6.6.4
Select the
required wavelength from the keyboard [e.g. 230nm- press key number 2, 3, 0 and
‘.’ Then ‘0’ and press key ENTER].
6.6.5
On screen the
wavelength drive will start moving to the selected wavelength [i.e.
600.0nm-230.0nm].
6.6.6
Clean the quartz
cells with purified water and then with blank solution.
6.6.7
Fill both the
quartz cells with blank solution. Clean cells with soft tissue paper.
6.6.8
Place the cuvettes
into the reference and sample compartment of the instrument and close the
shutter of the compartment.
6.6.9
Press Auto zero on
the keyboard “0.000” will appear on the screen.
6.6.10
Now the instrument
is ready for taking absorption at chosen wavelength for the sample.
6.6.11
Take out the cell
from the sample compartment of the instrument and rinse it with the sample
solution, and fill it to ¾ the height.
6.6.12
Clean the cell with
soft tissue paper and place it in the sample compartment of the instrument and
close the shutter.
6.6.13
The absorbance of
the sample will appear on the screen.
6.6.14
Print the
absorbance of the sample by pressing PRINT DATA key on the key board.
6.7
Wavelength
scan:
6.7.1
Set cursor to
wavelength scan, by pressing “ENTER”.
6.7.2
Now select
options,
6.7.2.1
ABS,
6.7.2.2
%T ,
6.7.2.3
SINGLE BEAM.
6.7.3
Press ENTER again
.TEST SET UP appears.
6.7.4
Press ENTER .One
more cursor will appear on the screen i.e. on START WL [nm] simultaneously
ENTER VALUE [190.0-1100.0] appears below the PARAM table.
6.7.5
ENTER the START WL
by pressing numerical keys.
NOTE: START WL means the highest wavelength in the wavelength scanning
range.
6.7.6
Then STOP WL appears ENTER VALUE [190.0-1100.0].
6.7.7
Select the lowest wavelength to be scanned by pressing
the numerical keys and ENTER.
NOTE: STOP WL [nm] is the lowest wavelength in the wavelength scanning
range.
6.7.8
Press ENTER UP - SCALE appears, select the maximum
absorbance to be kept as UP -SCALE.
6.7.9
Press ENTER to arrive ‘LO SCALE’. Select the lowest
absorbance on the or below the base line and feed by pressing numerical keys
[e.g. 0.000 the base line absorbance], normally it is kept – 0.100 i.e. below
the base line.
6.7.10
Press ENTER TO ‘SCAN SPEED’ [nm/min], select the options
out of 1 to 8 i.e. the options by pressing numerical keys.
6.7.11
Press ENTER to arrive to ‘INIT-DELAY’ [sec] i.e. the time
delay to start the scan, ENTER VALUE [0-9999] to feed the initial delay.
6.7.12
Press ENTER to go to ‘CYCLE TIME’ [sec], ENTER value from
0-9999 by pressing the numerical keys and then ENTER.
6.7.13
Press ENTER to DISPLAY FORMAT. Select options out of two
i.e.
6.7.13.1 SEQUENTIAL.
6.7.13.2 OVERLAY.
6.7.14
Press ENTER to test name.
6.7.15
Press ENTER to “ENTER” to change string.
6.7.16
Feed the name of the test scanning by operating the
CURSOR provided for SELECT – 0-10 Characters.
6.7.17
Enter to maximum 10 letters abbreviated or numerical to
be coded for TEST NAME.
6.7.18
Enter to INSTR SET UP and fix parameters.
6.7.19
BASELINE: select option i) system ii) user press ENTER to RESPONSE: select option i) Fast, ii)
Medium iii) Slow. (MEDIUM response for WL SCAN is desirable).
6.7.20
ENTER to LAMP CHARGE WL (nm) ENTER VALUE from
325.0-370.0.
6.7.21
ENTER the following like this (one after another).
6.7.21.1 VISLAMP-ON (select options i. ON ii. OFF).
6.7.21.2 UV LAMP-ON (select options i. ON ii. OFF).
NOTE: When we are doing any scanning in the UV region or VISIBLE region, we
need not keep the other lamp ON.
6.7.22
GRAPH PRINT: select option i. ON ii. OFF. Keep the
text print –ON when simultaneous Printing of graph is required during scanning.
6.7.23
TEXT PRINT: select option i) ON, ii) OFF. Keep the text print- ON
when simultaneous printing of absorbance at particular WL is desired at
particular intervals.
6.7.24
LIST INTERVAL: Enter the value (0.1-100.0) by pressing the
numerical keys.
6.8
Enter
to user baseline:
6.8.1
Keep the blank solution.
6.8.2
Again ENTER to START to measure USER BASE LINE. Make the
USER BASE LINE ‘AUTO ZERO’ for the WL range opted for two times.
6.8.3
FORWARD to get the graph i.e. WL against Absorbance.
6.8.4
Fill the sample cell with test solution, wipe with tissue
paper and fit in the sample compartment.
6.8.5
Start scanning and take prints if required. Trace peaks
on the scan by operating numerical keys.
6.8.6
Take out the quartz cells press ‘MAIN MENU’ on the
keyboard to get back the instrument to its original position.
6.8.7
Switch OFF the instrument after operation and clean the
quartz cells. For further operations, please refer manual.
7.0 REVISION LOG:
|
Revision No.
|
Effective Date
|
Reason
|
|
00
|
|
New
SOP
|
8.0 REFERENCES:
8.1
Not Applicable.
9.0 ANNEXURES:
9.1
Not Applicable.
10.0
ABBREVIATIONS:
|
Abbreviation
|
Expanded Form
|
|
SOP
|
Standard operating
procedure
|
|
&
|
And
|
|
No.
|
Number
|
|
Ltd.
|
Limited
|
|
Q.C
|
Quality control
|
|
QCEO
|
Quality control
equipment operation
|
|
F
|
Format
|
|
UV/VIS
|
Ultraviolet/ visible
|
|
λ
|
Lambda
|
|
nm
|
Nanometer
|
|
D2
|
Deuterium-2
|
|
ABS
|
Absorbance
|
|
%
|
Percentage
|
|
T
|
Transmittance
|
|
e.g.
|
For example
|