Matters that should be noted when determining substance content by spectrophotometry:
1, is the condition of color reaction
2, is the condition of measuring absorbance.
1, is the condition of color reaction
2, is the condition of measuring absorbance.
The conditions of the color reaction are
the amount of color developing agent, the color developing time, the elimination method of interfering substances, the acidity of the medium, the temperature of the solution when the color is developed. The conditions for measuring absorbance include the wavelength of the incident light that should be selected, the range of absorbance and the reference solution.
Ultraviolet spectrophotometer determination of trace iron color development agent can choose o-diazophene (phen) under the condition of pH=2 ~ 9, Fe2+ ions and o-diazophene produce stable orange-red complex, the reaction is as follows:
This complex lgK stability =21.3, molar absorption coefficient.
Before color development, Fe3+ ions are reduced to Fe2+ ions by hydroxylamine hydrochloride:
The determination should control the acidity of the solution at pH=5 or so is appropriate, the acidity is too high, the reaction is slow, the acidity is too low, Fe2+ ion hydrolysis, affecting the color.
Ultraviolet spectrophotometer determination of trace iron color development agent can choose o-diazophene (phen) under the condition of pH=2 ~ 9, Fe2+ ions and o-diazophene produce stable orange-red complex, the reaction is as follows:
This complex lgK stability =21.3, molar absorption coefficient.
Before color development, Fe3+ ions are reduced to Fe2+ ions by hydroxylamine hydrochloride:
The determination should control the acidity of the solution at pH=5 or so is appropriate, the acidity is too high, the reaction is slow, the acidity is too low, Fe2+ ion hydrolysis, affecting the color.
reagents and instruments
Instrument: UV-1500PC (Shanghai Meishe Instrument Co., LTD.)
Reagent:
1. 100 iron standard solution
2. 10 iron standard solution is prepared by dilution of 100
3. Hydroxylamine hydrochloride 10% solution
4. O-diazepine solution 0.15%
5. NaAc solution 1mol/L
Reagent:
1. 100 iron standard solution
2. 10 iron standard solution is prepared by dilution of 100
3. Hydroxylamine hydrochloride 10% solution
4. O-diazepine solution 0.15%
5. NaAc solution 1mol/L
Experimental steps
1. The choice of measurement wavelength
In 50ml volumeter bottle accurately add the following solution in order, 10 iron standard solution 5ml, 10% hydroxylamine hydrochloride 1ml, shake well, and then add 1mol/LNaAc solution 5ml, 0.15% o-diazophene 3ml. Dilute to scale with water. On the spectrophotometer, the absorbance between 440 and 580nm is determined by using a 1cm colorimetric dish with water as the
reference solution. And draw a graph to find out the wavelength of large absorption from the curve.
reference solution. And draw a graph to find out the wavelength of large absorption from the curve.
2. Color development agent dosage selection
Take 750ml volumetric bottles, each add 1ml10 iron standard solution, 1ml hydroxylamine hydrochloride, shake well. Then add 0.10, 0.30, 0.50, 0.80, 1.0, 2.0, 4.0ml ortho diazepine and 5ml NaAc solution, dilute to the scale with water, shake well, and leave for 10min. With water as the reference solution, the absorbance of each solution was determined at the selected wavelength. And draw, from the curve to find the appropriate amount of color and V.
3. The choice of color development time
In a 50ml volumetric bottle, add 1ml10 iron standard solution, 1ml hydroxylamine hydrochloride, and then add the suitable o-diazepine Vml and 5mlNaAc selected by the above, dilute to the scale with water, shake well. Immediately measure the absorbance at the selected wavelength with water as the reference in the colorimetric dish. Then measure the absorbance after 5, 10, 30, 60min in turn. Draw the graph and select the appropriate time for color reaction with o-diazepine.
4. Draw the standard curve
In 6 50ml volumetric bottles, add 0.0, 2.0, 4.0, 6.0, 8.0, 10.0ml of 10 iron standard solution, then add 1ml hydroxylamine hydrochloride, 2ml o-diazepine, 5mlNaAc solution, dilute with water to the scale, shake well. With water as the reference solution, the absorbance of each solution was determined under the selected wavelength, the selected appropriate color developer dosage and the appropriate color development time. Draw the standard graph.
Data recording and processing
1. Selection of measurement wavelength
| Wavelength /nm | 440 | 460 | 480 | 490 | 500 | 505 | 510 |
| Absorbance /A | 0.139 | 0.156 | 0.175 | 0.178 | 0.185 | 0.190 | 0.192 |
| Wavelength /nm | 515 | 520 | 530 | 540 | 560 | 580 | |
| Absorbance /A | 0.189 | 0.180 | 0.143 | 0.094 | 0.033 | 0.01 |
As shown in the figure above, the appropriate wavelength is =510nm
2. Selection of the amount of developing agent
Color developing agent volume V/mL
| Color developer volume V/mL | 0.1 | 0.3 | 0.5 | 0.8 | 1.0 | 2.0 | 4.0 |
| Absorbance /A | 0.044 | 0.048 | 0.048 | 0.043 | 0.055 | 0.048 | 0.054 |
From the figure above, the appropriate amount of indicator is V=1.0ml
3. The choice of color development time
| Colordevelopment time t/min | 2 | 5 | 10 | 30 |
| Absorbance /A | 0.055 | 0.056 | 0.056 | 0.058 |
From the above figure, the slope of the line K=1.0×10-4≈0, so it can be considered that the color development time has almost no effect on the absorbance.
4. Drawing of standard curve
Iron standard solution volume v/mL
| Iron standard solution volume v/mL | 2.0 | 4.0 | 6.0 | 8.0 | 10.0 | To bemeasured |
| Iron standard solution concentration /b | 0.4 | 0.8 | 1.2 | 1.6 | 2.0 | To bemeasured |
| Absorbance /A | 0.096 | 0.175 | 0.239 | 0.313 | 0.383 | 0.281 |
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