A spectrophotometer is an equipment that employs purer monochromatic light as incident light to measure the absorption of light by compounds and perform qualitative or quantitative analysis of substances. Digital spectrophotometers, UV spectrophotometers, fluorescence spectrophotometers, and other types of spectrophotometers are commonly used. During use, spectrophotometer measurement errors are common. How do these mistakes happen?
1.Polychromatic Light's Deviation from the Contrast Ear Law
Beer's law cannot be established unless the light emitted by humans is monochromatic. Even a spectrophotometer with a dual monochromator, however, can only produce nearly monochromatic light and cannot produce pure monochromatic light. It still has a narrow light passband and polychromatic light characteristics. Beer's law will deviate in either a positive or negative direction due to the polychromatic light. An ultraviolet spectrophotometer's spectral bandwidth is normally 1nm or 2nm, with a minimum adjustable slit of 0.1nm; a visible spectrophotometer's bandwidth is 6nm, or even more than ten nanometers.
The spectral bandwidth should be kept to a minimum. However, as the spectral resolution increases, the instrument's sensitivity decreases. As a result, when picking an instrument, the impact of various conditions should be examined. For example, when the concentration of the solution is low, the light is monochromatic, and the color of the light is purer, Beer's law can be roughly applied.
2.The effect of stray light
Stray light is defined as other wavelength components that enter the detector outside of the spectral bandwidth of the wavelength to be measured, and it is the primary source of error in spectrum measurement.
The reasons are the spectrophotometer's dispersion element, the mirror, the lens, and dust on the inner wall of the monochromator. Stray light has a greater influence at the edge wavelength of the spectrophotometer's working band due to the poor transmittance of the monochromator, the radiation intensity of the light source, and the sensitivity of the detector.
Stray light limit instrument analysis's upper limit can result in substantial measurement mistakes. In actual work, the absorbance of the sample is often measured at or near the absorption peak in quantitative analysis. If stray light is present at the analysis wavelength, the sample's transmittance rate is low, and most of the stray light is transmitted, causing the recorded absorbance to be lower than the true absorbance.
3.The impact of instrument noise on measurement Tool
Instrument noise is a good sign of instrument error. It represents the instrument's ability to prepare dilute solutions. It is an undesirable signal superimposed on the to-be-measured analytical signal. Scan the 100%T and 0%T lines to get the spectrophotometer's absolute noise level. The smaller measurement signal will be obscured if the instrument is noisy. In general, twice the noise is employed to describe the instrument's sensitivity.
4.Wavelength and absorbance precision
Each measured sample value should be measured at a given wavelength. The measured value will be erroneous if the wavelength error is significant.
Selection of measurement conditions
1. Selection of reference solution and solvent
To determine the absorbance of the sample, the spectrophotometer utilizes the light intensity of the reference cell as the light intensity of the person. Adjust the instrument so that the absorbance of the solution passing through the reference cell is zero, and then pass the same light through the sample. Because the absorbance truly reflects the concentration of the substance to be tested, the reference solution is critical.
Pure solvent or distilled water can be used as the reference solution if just the reaction product of the chemical is examined and the developer is absorbed. Use the developer solution as the reference solution if the developer has color and absorbs at the measurement wavelength. The developer and other reagents should be introduced in the same proportion as the sample.
Use the sample solution without the developer as the reference solution if the color of other components in the sample interferes with the measurement and the developer used has no color.
The correct solvent selection plays a vital role in improving the accuracy of the analysis. A high-purity solvent should be used to decrease the influence of impurities in the solvent; the solvent should not chemically react with the test material; the test substance must have a certain solubility in the solvent; and if the solvent itself does not absorb light within the measured wavelength range, When measuring absorption, the shortest available wavelength of common solvents should be used; if a volatile solvent is used, the absorption cell should be covered during the measurement.
2. Selection of test wavelength
When measuring the solution with a spectrophotometer, you must first select the appropriate measurement wavelength. The absorption curve of the tested solution is used to make the decision. In general, we select the greatest absorption wavelength as the measurement wavelength to improve sensitivity.
In some cases, the maximum absorption peak is very sharp, the absorption is too large, or there is interference nearby, and the other wavelengths in the absorption curve must be chosen for determination while ensuring a certain sensitivity (the curve is more Corresponding wavelength on the flat surface) to eliminate interference. Drawing the absorption curve is an effective tool and strategy for selecting the correct wavelength.
In general, there are two causes of spectrophotometer error: the error generated by the instrument's quality and manufacturing process, and the error induced by variable measurement conditions. Because there are so many different types of spectrophotometers on the market, the most important thing is to find one that is right for you.
I strongly recommend Drawer Scientific's spectrophotometers towards the end of the article. Digital spectrophotometers, UV spectrophotometers, fluorescence spectrophotometers, and other spectrophotometers are available from them. You can get things that are appropriate for your needs. If you have any questions regarding selecting a spectrophotometer, you may consult the engineers, who will be pleased to assist you.
