How to use the 722 spectrophotometer
First, the principle of measurement
The theoretical basis for the spectrophotometric measurement is the Bolang-Beier law: when the substance in the liquid is irradiated and excited by light, it produces an effect on light absorption. However, the absorption of light by matter is selective, and various substances have their own absorption spectra. Therefore, according to the law, when a bundle of monochromatic light passes through a rare color solution of a certain concentration range, the absorption degree A of the solution is proportional to the concentration c (g/l) of the solution or the thickness b (cm) of the liquid layer. Its law expression A=abc
(a is the proportionality factor). When the unit of c is mol/l, the proportional coefficient is represented by ε, and A = εbc is called the molar absorptivity. Its unit is L·mol-1·cm-1, which is the characteristic constant of a colored substance at a certain wavelength.
T (light transmittance) = I / I0 A (absorbance) = - lgT or A = K · C · L (the thickness of the cuvette)
When measured, incident light I,
When the light absorption coefficient and the optical path length of the solution are constant, the transmitted light varies depending on the concentration of the solution, that is, "K" is a constant. The thickness of the cuvette is constant, and "L" and "I0" are also fixed. As long as A is measured, "C" can be calculated.
On the meter of the spectrophotometer, one line is the light transmittance and one line is the absorbance. 》
Second, the use of 722 spectrophotometer
1. Set the sensitivity knob to “1†(the signal magnification is the smallest).
2. Turn on the power, the indicator light is on, the selector switch is set to “Tâ€, and the wavelength is adjusted to the test wavelength. The instrument is warmed up for 20 minutes.
3. Open the sample chamber (the light door is automatically closed) and adjust the light transmittance zero knob to make the digital display 000.0. (Adjust the 100% T knob), cover the sample chamber cover, place the cuvette holder in the distilled water calibration position, and let the photocell receive light. Adjust the light transmittance 100% knob to make the digital display 100.0. If the display is less than 100.0, the multiple of the microcurrent amplification can be appropriately increased. (The number of steps to increase the sensitivity should be repeated at the same time (3) adjust the "0" position of the light transmittance of the instrument), but try to make the magnification in the low range. This way the instrument will have higher stability.
4. After preheating, press (3) to adjust the position of “0†bit and “100%†of light transmittance several times in succession. After the instrument is stable, the instrument can perform the measurement work.
3. Measurement of absorbance "A" Place the selector switch in A. Adjust the absorbance zero adjustment knob so that the digital display is zero, then move the sample to be measured into the optical path, and the displayed value is the absorbance value of the sample to be tested.
4. Measurement of concentration c. Select the switch from “A†to “C†to put the sample with the calibration concentration into the optical path, adjust the concentration knob so that the digital display is the calibration value, and the sample to be tested can be read into the optical path. The concentration value of the sample to be tested.
Precautions:
1. After the measurement is completed, open the cover of the cassette, turn off the power switch, adjust the sensitivity knob to the lowest position, take out the cuvette, put the desiccant bag containing the silicone into the cassette, close the lid, and put the lid in the cuvette. The solution is poured into a beaker, washed with distilled water and placed back in the cuvette box.
2. The cuvettes supplied with each instrument should not be exchanged individually with the surface plates on other instruments.
Instructions
(1) Preheat the instrument Set the selector switch to “T†and turn on the power switch to warm up the instrument for 20 minutes. In order to prevent the phototube from fatigue, do not continue to illuminate. When preheating the instrument and when not measuring, the sample chamber cover should be opened to cut off the optical path.
(2) Selected wavelengths According to the experimental requirements, turn the wavelength handwheel to the desired monochromatic wavelength.
(3) Fixed sensitivity file In the case that the blank solution can be adjusted to “100%â€, use the lower sensitivity as much as possible. When using it, first adjust to “1†block. When the sensitivity is not enough, then gradually increase. high. However, after the shift changes sensitivity, “0%†and “100%†must be recalibrated. Select the sensitivity and do not change during the experiment.
(4) Adjust T=0% Gently turn the “0%†knob to display the number as “00.0†(the sample chamber is open at this time).
(5) Adjust T=100% Put the cuvette containing distilled water (or blank solution, or pure solvent) into the first grid in the cuvette holder, and align the light path, gently light the sample chamber cover Cover and adjust the transmittance “100%†knob so that the digital display is exactly “100.0â€.
(6) Measurement of absorbance Place the selector switch in “Aâ€, cover the sample chamber cover, place the blank liquid in the light path, and adjust the absorbance adjustment knob so that the number is displayed as “.000â€. Place the cuvette containing the solution to be tested into other cells in the cuvette holder, cover the sample chamber cover, and gently pull the sample holder handle to make the solution to be tested enter the optical path. That is, the absorbance value of the solution to be tested. After reading, open the sample chamber cover and cut off the light path.
The above measurement operation was repeated 1-2 times, and the corresponding absorbance values ​​were read and averaged.
(7) Determination of concentration Select the switch to “C†by “Aâ€, put the sample with the calibration concentration into the optical path, adjust the concentration knob so that the digital display is the calibration value, and put the sample to be measured into the optical path. The displayed value is the concentration value of the solution to be tested.
(8) After the shutdown test is completed, turn off the power, remove the cuvette, and wipe the cuvette holder with a soft paper.
The theoretical basis for the spectrophotometric measurement is the Bolang-Beier law: when the substance in the liquid is irradiated and excited by light, it produces an effect on light absorption. However, the absorption of light by matter is selective, and various substances have their own absorption spectra. Therefore, according to the law, when a bundle of monochromatic light passes through a rare color solution of a certain concentration range, the absorption degree A of the solution is proportional to the concentration c (g/l) of the solution or the thickness b (cm) of the liquid layer. Its law expression A=abc
(a is the proportionality factor). When the unit of c is mol/l, the proportional coefficient is represented by ε, and A = εbc is called the molar absorptivity. Its unit is L·mol-1·cm-1, which is the characteristic constant of a colored substance at a certain wavelength.
T (light transmittance) = I / I0 A (absorbance) = - lgT or A = K · C · L (the thickness of the cuvette)
When measured, incident light I,
When the light absorption coefficient and the optical path length of the solution are constant, the transmitted light varies depending on the concentration of the solution, that is, "K" is a constant. The thickness of the cuvette is constant, and "L" and "I0" are also fixed. As long as A is measured, "C" can be calculated.
On the meter of the spectrophotometer, one line is the light transmittance and one line is the absorbance. 》
Second, the use of 722 spectrophotometer
1. Set the sensitivity knob to “1†(the signal magnification is the smallest).
2. Turn on the power, the indicator light is on, the selector switch is set to “Tâ€, and the wavelength is adjusted to the test wavelength. The instrument is warmed up for 20 minutes.
3. Open the sample chamber (the light door is automatically closed) and adjust the light transmittance zero knob to make the digital display 000.0. (Adjust the 100% T knob), cover the sample chamber cover, place the cuvette holder in the distilled water calibration position, and let the photocell receive light. Adjust the light transmittance 100% knob to make the digital display 100.0. If the display is less than 100.0, the multiple of the microcurrent amplification can be appropriately increased. (The number of steps to increase the sensitivity should be repeated at the same time (3) adjust the "0" position of the light transmittance of the instrument), but try to make the magnification in the low range. This way the instrument will have higher stability.
4. After preheating, press (3) to adjust the position of “0†bit and “100%†of light transmittance several times in succession. After the instrument is stable, the instrument can perform the measurement work.
3. Measurement of absorbance "A" Place the selector switch in A. Adjust the absorbance zero adjustment knob so that the digital display is zero, then move the sample to be measured into the optical path, and the displayed value is the absorbance value of the sample to be tested.
4. Measurement of concentration c. Select the switch from “A†to “C†to put the sample with the calibration concentration into the optical path, adjust the concentration knob so that the digital display is the calibration value, and the sample to be tested can be read into the optical path. The concentration value of the sample to be tested.
Precautions:
1. After the measurement is completed, open the cover of the cassette, turn off the power switch, adjust the sensitivity knob to the lowest position, take out the cuvette, put the desiccant bag containing the silicone into the cassette, close the lid, and put the lid in the cuvette. The solution is poured into a beaker, washed with distilled water and placed back in the cuvette box.
2. The cuvettes supplied with each instrument should not be exchanged individually with the surface plates on other instruments.
Instructions
(1) Preheat the instrument Set the selector switch to “T†and turn on the power switch to warm up the instrument for 20 minutes. In order to prevent the phototube from fatigue, do not continue to illuminate. When preheating the instrument and when not measuring, the sample chamber cover should be opened to cut off the optical path.
(2) Selected wavelengths According to the experimental requirements, turn the wavelength handwheel to the desired monochromatic wavelength.
(3) Fixed sensitivity file In the case that the blank solution can be adjusted to “100%â€, use the lower sensitivity as much as possible. When using it, first adjust to “1†block. When the sensitivity is not enough, then gradually increase. high. However, after the shift changes sensitivity, “0%†and “100%†must be recalibrated. Select the sensitivity and do not change during the experiment.
(4) Adjust T=0% Gently turn the “0%†knob to display the number as “00.0†(the sample chamber is open at this time).
(5) Adjust T=100% Put the cuvette containing distilled water (or blank solution, or pure solvent) into the first grid in the cuvette holder, and align the light path, gently light the sample chamber cover Cover and adjust the transmittance “100%†knob so that the digital display is exactly “100.0â€.
(6) Measurement of absorbance Place the selector switch in “Aâ€, cover the sample chamber cover, place the blank liquid in the light path, and adjust the absorbance adjustment knob so that the number is displayed as “.000â€. Place the cuvette containing the solution to be tested into other cells in the cuvette holder, cover the sample chamber cover, and gently pull the sample holder handle to make the solution to be tested enter the optical path. That is, the absorbance value of the solution to be tested. After reading, open the sample chamber cover and cut off the light path.
The above measurement operation was repeated 1-2 times, and the corresponding absorbance values ​​were read and averaged.
(7) Determination of concentration Select the switch to “C†by “Aâ€, put the sample with the calibration concentration into the optical path, adjust the concentration knob so that the digital display is the calibration value, and put the sample to be measured into the optical path. The displayed value is the concentration value of the solution to be tested.
(8) After the shutdown test is completed, turn off the power, remove the cuvette, and wipe the cuvette holder with a soft paper.
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