scinco Mega-900 UV-Vis User manual

0
5.
Mega-900 UV-Vis Spectrophotometer
Instruction Manual
Janurary 2016

Mega-900 SCINCO
Introduction
Thanks for your purchase of the SCINCO CO., LTD. Mega-900 spectrophotometer.
UV-Visible Spectrophotometer is a well-accepted, documented technique with many
applications. The technique is extensively used for the analysis of agricultural
products and is widely used in public health, environmental protection, life sciences
industries and many other organic and biochemical applications.
For obtaining maximum performance and trouble-free operation, we give you the
following suggestions:
◆Upon receiving your instrument, please check all the items listed in the
packing list are included and install the instrument according to the instruction
manual.
◆The spectrophotometer is a precise electronic instrument. Please read the
instruction manual carefully before operation. If the equipment is used in a
manner not specified by the manufacturer, the protection provided by the
equipment may be impaired.
◆Avoid collision and handle the instrument carefully during transportation.
◆If the tested solution is harmful to humans, please wear gloves or use other
protection methods.
◆Please keep the accessories in a safe place for the use in future maintenance.
For more information, please visit www.scinco.com

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Contents
Contents.....................................................................................................................I
Chapter 1 Instrument Introduction..........................................................................1
1.1 Components inspection....................................................................................1
1.2 UV/Vis spectrophotometer and method............................................................1
1.2.1 UV/Vis spectrophotometer method................................................................1
1.2.2 UV/Vis spectrophotometer ............................................................................1
1.2.3 Condition of measure method .......................................................................2
1.3 Components description ..................................................................................3
1.3.1 Front view .....................................................................................................3
1.3.2 Rear view......................................................................................................4
1.3.3 Side view ......................................................................................................4
1.3.4 Sample cell holder.........................................................................................5
1.4 Specifications...................................................................................................6
Chapter 2 Installation...............................................................................................7
2.1 Installation location.........................................................................................7
2.2 Installation......................................................................................................7
2.3 Start ...............................................................................................................8
2.4 Performance inspection in the installation process.......................................10
2.4.1 The accuracy and reproducibility of wavelength..........................................10
2.4.2 Flatness of the baseline ..............................................................................11
Chapter 3 Instrument operation ............................................................................13
3.1 About double beam measurement................................................................13
3.1.1 What’s the double beam..............................................................................13
3.1.2 How to use double beam instrument...........................................................14
3.2 Application examples ...................................................................................15
3.2.1 Photometric measurement example............................................................15
3.2.2 Spectrum measurement example................................................................16
3.2.3 Quantitative measurement example............................................................18

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Chapter 4 Optional accessories ............................................................................23
4.1 CH19-1 Thermostatic cell holder....................................................................23
4.2 DS19-1 Angle-changeable solid sample holder..............................................24
4.3 IS19-1 Integrating sphere...............................................................................25
4.4 LS19-1 Long pathlength cell holder................................................................26
4.5 MH19-1 Micro cell holder ...............................................................................27
4.6 MH19-2 Ultra-micro cell holder.......................................................................28
4.7 MR19－1 Specular reflection accessory.........................................................29
4.8 MS19-1 Manual 8-cell holder .........................................................................30
4.9 PS19-2 Sipper pump......................................................................................31
4.10 S19-1 Solid sample holder............................................................................33
4.11 SS19-1 Short pathlength cell holder..............................................................34
4.12 TR19-1 Test tube cell holder.........................................................................35
Chapter 5 Maintenance ..........................................................................................36
5.1 Points for attention:........................................................................................36
5.2 Daily care.......................................................................................................36
5.3 Trouble shooting and maintenance................................................................36
Appendix 1 Light source replacement..................................................................39
Replace the tungsten lamp...................................................................................39
Replace the Deuterium lamp ................................................................................40
Appendix 2 Fuse replacement...............................................................................42

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Chapter 1 Instrument Introduction
1.1 Components inspection
After the instrument has been unpacked, please check up carefully the parts provided
against the packing list for acceptance. In the event of error or damage, contact the
sales agent immediately for prompt solution. A complete set of the instrument is
according to the packing list in the instrument package. The packing list shall be taken as
the criterion. The company keeps the right to revise.
1.2 UV/Vis spectrophotometer and method
1.2.1 UV/Vis spectrophotometer method
UV/Vis spectrophotometer method is an instrument analysis method that
measures the molecule’s absorbance of radiation in ultraviolet and visible
spectrum. The absorption of molecule corresponds to the excitation of valence
electrons and electrons in the molecular orbital between different electronic energy
level, this method can be used for qualitative and quantitative analysis of organic
and inorganic compounds.
Lambert-Beer’s law is the basic law of radiation absorbance; it’s the foundation of
spectrophotometer analysis method. The absorbance (A) of solution is directly
proportional to the path length (b) and the concentration (c) when the wavelength
of the incidence light is fixed. Lambert-Beer’s law states that
ClA 

, where

is the constant of proportionality, called the absorbtivity.
1.2.2 UV/Vis spectrophotometer
UV/Vis spectrophotometer is the general analysis instrument which is basic on
the principle of UV/Vis spectrophotometer method. UV/Vis spectrophotometer can
be classified by single beam spectrophotometer, double beam spectrophotometer,
and double wavelength spectrophotometer according to the difference of optical
structure.
All kinds of UV/Vis spectrophotometer is composed of five basic component, they
are light source, monochrometer, cell holder, detector and signal processing
system.

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1.2.3 Condition of measure method
The choice of measuring wavelength
Normally taking the wavelength which has the most absorbance as the
measuring wavelength (
max

) to attain the highest analysis sensitivity is called
most absorbance principle. And the absorbance will change little along with
wavelength’s changing around the
max

, so we can get a better precision. But we
would like to take the low sensitivity measuring wavelength as the absorption
peak wavelength when measuring high concentration compounds, in order to
guarantee the corrective curve has enough linear range. If the absorption peak in
the wavelength
max

is too thin, we can adopt a little lower sensitivity wavelength
to measure on the condition that satisfies the need of analysis sensitivity, which is
to decrease the deviation of Beer’s Law.
The choice of absorbance range
Because of the unstable of light source, inaccuracy of reading, or alternation of
lab condition in the measurement, any photometer has some measuring error.
Because the transmittance T in the absorbance law is negative logarithm to the
concentration C, from the curve of negative logarithm we can see, the relative
error of the same transmittance in different concentration is different. The relative
error is very big when the concentration is too high or too low, so we should
choose the proper absorbance range to decrease the relative error of the result.
In practice, we can adjust the concentration of the solution or the length of the
cell to make the absorbance in the require range.
The choice of bandwidth
Bandwidth will affect the sensitivity of measurement and linear range of
correction curve. If the bandwidth is too wide, the monochromatic light of incidence
will be less, the correction curve will deviate from Beer’s Law, and the sensitivity
will be lower. If the bandwidth is too thin, the gain of the instrument will be very
high, the noise will be increase, and it’s not good for measurement. Measure the
deviation of absorbance along with the bandwidth so as to choose the proper one.
The absorbance will not change if the bandwidth is in a range, but the absorbance
will decrease if the bandwidth is as wide as some level. So the widest bandwidth
which is not decrease the absorbance is the proper one we should choose.

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1.3 Components description
When unpacking, the instrument is present as shown in Figure 1.1, next is the
detailed description of all components of the instrument.
Figure 1.1 Instrument overview
1.3.1 Front view
The front view is shown as Figure 1.2, the name and function of all
components is described below:
Figure 1.2 Front view
①Instrument cover
②Front baffle of sample compartment—standard front baffle for sample
compartment. It should be replaced when using some accessories.
③Chassis—the steel made chassis can greatly reduce vibration, and ensures
the credibility of the instrument.

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④Instrument label
⑤Sample compartment cover—Open the cover to gain access to the sample
compartment. The sample compartment is used to hold sample, it can be
replaced by manual 8-cell holder, long path length cell holder, and fixed cell
holder and so on.
1.3.2 Rear view
The rear view is shown as Figure 1.3, the name and function of all components
is described below:
Figure 1.3 Rear view
①Fan
②Foot packing—Shockproof rubber washer is used.
③RS-232 interface—Communicate with PC.
④Power supply—Connect to power supply: AC 220V, 50Hz.
⑤Fuse 1—Can be replaced (refer to “Appendix 3 fuse replacement”).
⑥Fuse 2—Can be replaced (refer to “Appendix 3 fuse replacement”).
⑦Instrument nameplate
1.3.3 Side view
The side view is shown as Figure 1.4, the name and function of all components
is described below:

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A Left side view B Right side view
Figure 1.4 Side view
①Power switch
②Light source house cover—Open the cover to gain access to the light source
house, where you can replace the deuterium lamp and tungsten lamp (refer
to “Appendix 1 light source replacement”).
1.3.4 Sample cell holder
The sample cell holder is configured according to the user’s requirements. The
general configurations are fixed cell holder, manual 8-cell holder, and long path
length cell holder. We can also provide you multiple kinds of sample cell holders
for your selection. Please refer to “chapter four Accessories” for the detailed
information.
The following figure 1.5 shows the fixed cell holder which is used as standard
configuration. This kind of sample cell holder is the most general structure, there
are two standard cell holders, and one is reference cell holder for blank sample,
while another is sample cell holder for measured sample.
Figure 1.5 Fixed cell holders

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1.4 Specifications
Figure 1-3
Model
Mega-900
Wavelength range
190nm ~ 90nm
Spectral bandwidth
Continuous slit 0.1 ~ 5.0 nm with 0.1 nm interval
Setting wavelength
The start and end wavelength can be set by the span
of 1nm. The goto-wavelength operation can be set by
the span of 0.1nm.
Wavelength accuracy
±0.3nm (with built-in automatic correction)
Wavelength
reproducibility
≤0.15 nm
Wavelength of
changing lamp
Set the wavelength of changing lamp within the
scope of 322-392nm at 1nm interval
Photometric system
Double beam, dynamic feedback direct ratio
recording system
Photometric scope
-4～4 Abs
Recording scope
Absorbance:-9.999～9.999 Abs
Transmittance:-999.9～999.9 %T
Photometric Accuracy
±0.3 %T (0~100%T)
Photometric
reproducibility
≤0.1 %T (0~100%T)
Stray light
≤0.01%T（220nm NaI）
≤0.05%T（340nm NaNO2）
Baseline flatness
±0.001 Abs ( scan with 850~200nm)
Stability
≤0.0004 Abs/h (2hr warm-up, kinetic scan at 500nm)
Noise
≤0.0004 Abs (500nm)
Note:
1. The above specifications are got at 2nm spectrum bandwidth. The specifications
are different at other spectrum bandwidths.
2. The above specifications are subject to change without notice.

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Chapter 2 Installation
2.1 Installation location
Mega-900 double-beam UV/Vis spectrophotometer (here in after referred as Mega-
900) is a versatile high-precision laboratory instrument. For enhance of its performance
and its usable longevity, the instrument should be installed in a suitable location with
the following requirements:
①Avoid high temperature and high humidity environment. An environmental
temperature should be 15℃~35℃. The relative in-door humidity should be 45~85%.
The relative in-door humidity should be lower than 70% if the in-door temperature is
over 30℃
②The instrument should be placed on a sturdy workbench that is free of a strong
vibration source.
③The location room should be free from a strong electromagnetic interference
source or harmful or poisonous gases.
④Power Supply: 220V±10%, 50Hz±1Hz. A voltage regulator (500VA).
Ensure satisfactory earth connection for the instrument.
2.2 Installation
MEGA-900 is composed of the three major components:
①A UV/Vis spectrophotometer main unit.
②An IBM-compatible Personal Computer completed with a monitor and a keyboard.
For the computer outfitted by user himself, there is the special control card, being
offered in the packing accessories, which is responsible for controlling the
spectrophotometer unit and should be plugged in the socket in the computer cabinet.
③Software: The operate software is LabPro Scan. You should setup the software in
the computer, about the detailed information you can find in the manual of LabPro
Scan. Please pay attention to the SN of the software LabPro Scan on the package of
software.
④A printer.
They are connected as shown Figure 2.1:

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Figure 2.1
Connect the instrument’s RS 232 communication port to the PC with RS 232 cable.
Connect the printer’s parallel port or USB port to the PC with communication cable,
and setup the printer driver in the PC (the detailed information refer to the printer
manual)
Both ends of all the connecting cables should be plugged tight to the sockets to
prevent poor connection. Only they will be allowed to connect the power cords of
instrument, PC, and printer to the power supply connector. (Note: make sure that the
rating of the power source be an AC of 220 V with 50 HZ.)
Note: The spectrophotometer is a precision electronic instrument. Don’t pull
out or insert the power cable or tie cable when the instrument is on, otherwise it
will destroy the instrument and the computer.
2.3 Start
Be sure to there is nothing to block in the sample cell.
First turn on the power for printer, then the power for computer. And then turn on the
power for main unit. The computer will start OS automatically. Select LabPro Scan
software in the Start menu, carry through Self Checking if it detects the instrument,

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the interface is shown as figure 2.2. If the software shown: “Warning: Unable to
communicate with main unit……” before the self checking, it means the software can’t
connect with the instrument, please check if the instrument and PC communication
connect well or not, if the instrument is power on or not, and then restart the
instrument, choose “Retry” menu.
Figure2.2 Initialization
After self checking, enter main working program. The interface is shown as figure 2.3.
Then you can operate on the instrument. The measurement can only be done for an
interval of preheating lasting 60 minutes after switching on the instrument in order to
insure the precision of result. During this time of initialization we advise you being
familiar with the operation of the instrument and software by instruction manual on the
back. If you need the instruction manual of the LabPro Scan software please refer to
the software manual and help document of the software menu.
After the measurement, first turn off the power supply for mail unit, then the power
supply for printer, at last the power supply for computer and other equipments (such
as voltage regulator).
Notice:
Please check for obstacle in the sample compartment if error of
initialization after power on, and restart the instrument. If can’t still
initialize normally, please contact with the supplier.

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Figure 2.3 LabPro Scan interface
2.4 Performance inspection in the installation process
An overall check-up of the major performances of the instrument should be made
after its installation. During the course of use, periodical inspection should also be
made. The checkup can only be done for an interval of preheating last 60 minutes
after switching on the instrument. The testing method should be referred to the
instruction manual below.
2.4.1 The accuracy and reproducibility of wavelength
The wavelength accuracy of the instrument is ±0.3nm ，whereas its
reproducibility is 0.15 nm (Please refer to the Chapter 1.4 Specification). The
checkup should be made in the following manner by using the two characteristic
spectrum lines of the deuterium lamp.
Setting the parameters by using the spectrum scan function (refer to LabPro
Scan software manual) as shown in table 2-1 and table 2-2:
Table 2-1（656.1nm characteristic spectrum）
Scan parameter
Instruction
Photometric
Es
Energy for sample light
Scan speed
Medium
Sample interval
Automatic
1nm
Energy condition
D2 lamp, gain 2 /3
Reference
Wavelength range
660 ~ 650 nm
Vertical coordinate
range
0 ~ 100
Notice:
There may be some time drift of instrument because of the long time preheating,
so please to do the wavelength correction and dark current correction before
measurement in order to get the most precision result. The detailed instruction
should be refer to the LabPro Scan software manual.

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Instrument parameter
Instruction
Spectrum bandwidth
2.0nm
Can’t be changed if fixed
Table 2-2（486.0nm characteristic spectrum）
Scan parameter
Instruction
Photometric
Es
Energy of sample light
Scan speed
Medium
Sample interval
Automatic
1nm
Energy condition
D2 lamp, gain 2/3
Reference
Wavelength range
490 ~ 480 nm
Vertical coordinate
range
0 ~ 100
Instrument parameter
Instruction
Spectrum bandwidth
2.0nm
Can’t be changed if fixed
Scan three times and pick the peaks. Note the three peaks and calculate the
accuracy of wavelength.
Calculate the mean value of wavelength

according to the formula (2-1)
Formula:



n
ii
n1
1

…………………………（2-1）
i

—— measured value of wavelength
n
—— measured times, here n=3
The difference value between the mean value and standard value is the
wavelength accuracy. The difference value between the maximum and the
minimum is the wavelength reproducibility.
2.4.2 Flatness of the baseline
Testing method: Check that the sample beam is blank (empty with no solution).
Then set the measuring parameters by using the spectrum scan function (refer
to the LabPro Scan software manual) as shown in table 2-3
Table 2-3（measure the flatness of baseline）
Scan parameter
Instruction
Photometric mode
Abs
Energy of sample light
Scan speed
Slow
Sample interval
Automatic
1nm
Wavelength range
900 ~ 190 nm
Vertical coordinate
range
-0.01 ~ 0.01
Instrument parameter
Instruction
Spectrum bandwidth
2.0nm
Can’t be changed if fixed

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Calibrate the baseline first, and then set the range of wavelength as 850-200nm,
and start for spectrum scanning. Read the absorbance of the spectrogram by the
graph reading function. The largest absorbance should conform to the
requirement of the flatness of the baseline. (A sudden change at the wavelength
of lamp changeover is permitted.)

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Chapter 3 Instrument operation
All of the operation of Mega-900 is almost performed by LabPro Scan software after
connecting with PC. Please read the LabPro Scan software manual in detail before operation
on it. This chapter will introduce the character of double beam measurement, and the
process of operation of Mega-900 by three examples.
3.1 About double beam measurement
3.1.1 What’s the double beam
Double beam UV/Vis spectrophotometer takes two light beams who’s energy
are the same to measure sample and reference respectively at the same time as
it’s optical principle (as figure 3.1 shown, R means reference light beam and S
means sample light beam), increase the stability and precision of the instrument
by the stability of real time feedback compensation measure system of the
reference beam.
The most difference between double beam, split beam and single beam on
structure is that the double beam instrument has two light beams in the sample
compartment, and reference and sample can be measured at the same time (as
figure 3.2 shown).

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Figure 3.1 Optical Principle
Figure 3.2 Sample cell holders
3.1.2 How to use double beam instrument
The difference between double beam and single beam (or split beam) on the
application is that to single beam you should put the blank and sample into the
sample cell holder to measure at twice when zero and measurement, while to
double beam just once.
To double beam instrument you just have to put the black block into sample light
beam cell holder to do the dark current correction.
You can familiar with the operation of Mega-900 by the following several
application examples.
Reference cell holder
Sample cell holder

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3.2 Application examples
3.2.1 Photometric measurement example
Vitamin B1 can promote the metabolizability of carbohydrate and fat, provide
energy for nervous tissue, prevent nervous tissue from atrophy and
degeneration, prevent and cure beriberi. The quality determination method of
Vitamin B1 for pharmaceutical factory is to measure the absorbance at 400nm;
it will be the conforming unit if the value is less than 0.020.
We need to adopt photometric measurement function to determinate the quality
of Vitamin B1 by LabPro Scan software, the detailed operation steps are as
follows:
①Prepare reagent
Weight 1g Vitamin B1 in a precision microbalance. Put the 1g of Vitamin
into a 10ml volumetric flask. Add adequate distilled water to dissolve the
Vitamin B1 to be the sample solution. Get two clean matched quartz cells
to fill with distilled water as the reference solution.
②Power on the instrument
Power on the PC which connect with the Mega-900, enter into the
Windows operate system. Be sure to there is nothing to block in the
sample cell holder, and power on the instrument, start LabPro Scan
software, carry through self checking. After self checking, enter main
working program. The measurement can only be done for an interval of
preheating lasting 60 minutes after switching on the instrument in order to
insure the precision of result.
③Setting parameters
Select “Photometry” function and enter into the parameter setting interface,
and set the parameters as follows:
Table 3-2 (Photometric measurement example parameter settings)
Measurement parameters
Instruction
Photometric mode
Abs
Measurement
wavelength
400nm
Instrument parameter
Instruction
Spectral bandwidth
2.0nm
Can’t be changed if fixed
④Dark current correction
Put the black block into the sample cell holder of sample light beam, to do
the dark current correction by the dark current correction function of
LabPro Scan software. Take out the black block after correction.
Notice：Dark current correction can eliminate some noise of the

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instrument, and insure the precision of result. You don’t have to do
it when at every measurement, just when the environment has been
changed, such as the temperature changed to much, the location
has been changed, measuring high absorbance sample and so on.
(Advise you doing the dark current correction after preheating and
before measurement, to insure the precision of the result.)
⑤Zero
Put the two matched quartz cells which are filled with distilled water into
the sample light beam cell holder and reference light beam cell holder,
cover the sample compartment cover, press “Zero” button of the LabPro
Scan software to zero, the absorbance shown on the top of the main
interface should be 0.000 Abs or nearby.
⑥Measure the sample
Take the cell out of sample light beam cell holder, and throw out distilled
water, wash the cell at least 3 times by Vitamin B1 solution and then filled
with sample solution. Put the cell which is filled with sample solution into
sample light beam cell holder, cover the sample compartment cover,
press “Start” button of the LabPro Scan software to finish the
measurement. After finishing measurement one time, press “Start” again
to measure the next data. We can get the result by the averaging of
several measurements.
⑦Analyze the result
You can save the result as special file format of LabPro Scan software
for checking and printing later.
3.2.2 Spectrum measurement example
Vitamin B12’s characteristic absorption peak is at 361nm, so the quality
determination method of Vitamin B12 is to measure the absorbance at 361nm,
next will take Vitamin B12 for example to show you how to use the spectrum
measurement function.
The detailed operation step of determination the absorption peak of Vitamin B12
by Mega-900 is as follows:
①Prepare reagent
Fetch Vitamin B12 solution, mixed it with distilled water according to the
proportion of 1:10 to get 10ml diluted solution. Get two clean matched
quartz cells to fill with distilled water as the reference solution.
②Power on the instrument
Power on the PC which connect with the Mega-900, enter into the
Windows operate system. Be sure to there is nothing to block in the