Since the most widely used detector for high performance liquid chromatography is a UV detector, it is used to detect samples with UV absorption. The UV detector is highly sensitive, does not destroy the sample, can be used in series with other detectors, and can also be used for preparation; it is insensitive to temperature and mobile phase flow fluctuations and can be used for gradient elution. However, only compounds having a π-π or p-π conjugated structure can be detected.
The cutoff wavelength of the solvent should be considered when using a UV detector. For example, when the detection wavelength is 220 nm, only a solvent smaller than the cutoff wavelength, such as n-pentane, water, methanol, acetonitrile or the like, may be used, and a solvent having a cutoff wavelength of less than 220 nm, such as dichloromethane, chloroform or the like, cannot be used.
The UV cut-off wavelength is defined as: "Air as a reference, the solvent absorbs the absorption wavelength equivalent to the reference in the 1 cm absorption cell".
When a solvent accounts for a small proportion of the mobile phase, it should be determined on a case-by-case basis. For example, a solution having a concentration of 10% (volume fraction) or less will detect only a background absorption of 0.1 AU (absorption unit) near the cut-off wavelength. In addition to the resulting increase in noise levels, linear operating range reduction and stability degradation, most cases are acceptable.
It should be noted that there may be significant differences in the spectral characteristics of different batches of solvents from different LC manufacturers. Due to the difference in the type and content of impurities, the UV cutoff wavelength and absorption coefficient will be different. The difference in the amount of oxygen dissolved in the air can clearly produce different baseline noise. Therefore, in addition to using solvents that meet chromatographic standards as much as possible, the solvent of the liquid chromatograph should be carefully filtered before use, degassed by ultrasonication, or even treated by blowing gas to remove impurities and volatile dissolved gases as much as possible.