(1). Quantitative analysis by gas chromatography/mass spectrometry (GC-MS)
The total ion chromatogram or mass chromatogram obtained by GC-MS has a peak area proportional to the content of the corresponding component. If a certain component is quantitatively determined, the normalization method in the chromatographic method can be used. Different methods such as external standard method and internal standard method are carried out. At this time, the GC-MS method can be understood as a detector of a chromatograph. The rest are the same as the chromatography. Unlike chromatographic quantification, GC-MS can be quantified using total ion chromatograms and quantified using mass chromatograms. This will minimize the interference of other components.
In order to improve detection sensitivity and reduce interference from other components, mass spectrometers often use selective ion scanning in GC-MS quantitative analysis. For the component to be tested, one or several characteristic ions may be selected, and the adjacent components do not have these ions. The chromatogram thus obtained has no interference in the component to be tested and has high sensitivity. Quantitative analysis was performed using chromatograms obtained from selected ions, and the specific analytical method was similar to the mass chromatogram. However, its sensitivity is higher than using a mass chromatogram, which is commonly used in GC-MS quantitative analysis.
(2). Liquid chromatography/mass spectrometry (LC/MS) analysis method
The mass spectrum obtained by LC-MS analysis is too simple, and the structural information is too small. It is difficult to conduct qualitative analysis. It mainly depends on the standard sample. For most samples, the retention time is the same, the product ion spectrum is the same, and the qualitative analysis can be obtained, except for a few isomers.
Quantitative analysis by LC-MS is the same as that of ordinary liquid chromatography. That is, quantification is performed by the peak area of the chromatogram and the correction factor (or standard). However, due to problems with chromatographic separation, a chromatographic peak may contain several different components, causing errors in quantitative analysis. Therefore, for LC-MS quantitative analysis, the total ion chromatogram is not used, but the mass chromatogram or multi-ion monitoring chromatogram obtained by using the characteristic ions corresponding to the component to be tested, at this time, the unrelated components will be no peak, which can reduce the mutual interference between components. LC-MS often analyzes samples with very complicated systems. There are a large number of interfering components with the same retention time and the same molecular weight. To eliminate interference, the best way to quantify LC-MS is to use multiple reaction monitoring (MRM) techniques for tandem mass spectrometry. That is, a sub-ion spectrum is prepared for the component to be tested of mass m1, and a characteristic ion m2 is selected from the sub-ion spectrum. When the sample is formally analyzed, the first-stage mass spectrum selects m1, and after collision activation, the second-stage mass spectrum selects m2. Only ions with the characteristic mass of both m1 and m2 are recorded. The chromatogram thus obtained was selected three times: LC selected the retention time of the component, the first stage MS selected m1, and the second stage MS selected m2, so that the obtained peak can be considered to have no interference. Then, according to the peak area of the chromatogram, the external standard method or the internal standard method is used for quantitative analysis. This method is suitable for the analysis of samples with low composition of the tested components, complex system components and severe interference.
5. Spectrum analysis
5.1. Spectrum and mass spectrometer
The mass spectrum refers to a spectrum composed of the mass-to-charge ratio of ions and their relative abundance after ionization of a compound molecule. It is conventionally called a bar graph.
The information expressed bymass spectrometry is simple and intuitive, but the expression of mass-to-charge ratio and abundance is not detailed and accurate, and it is insufficient when the spectrum is analyzed. Therefore, mass spectrometry is also often expressed in tabular form.
5.2.Spectrum analysis process
(1). Analyze the molecular ion region
a) Mark the mass-to-charge ratio of each peak, paying particular attention to the peaks in the high mass-to-charge ratio region.
b) Identify molecular ion peaks.
c) Analyze the relative intensity ratio of the isotope peak clusters and the Dm values between the peaks and the peaks to determine whether the compounds contain elements such as C1, Br, S, and Si, and elements such as F, P, and I, which are not isotopic.
d) Deriving the formula and calculating the degree of unsaturation. The molecular formula is calculated from the exact molecular weight measured by a high resolution mass spectrometer or from the relative intensities of the isotopic peak clusters.
e) Understand the molecular structure information from the relative intensities of the molecular ion peaks. The relative intensity of the molecular ion peak is determined by the structure of the molecule, and the structural stability is large and the relative intensity is large.
(2). Analyze fragment ions
a) Understand the possible structural information from the characteristic ion peaks and the missing neutral fragments.
b) Comprehensive analysis of all the information obtained above, combined with molecular formula and unsaturation, to propose the possible structure of the compound.
c) Analyze the cleavage excitation of the possible structure deduced to see if it matches the mass spectrum, determine its structure, and further interpret the mass spectrum, either in comparison with a standard spectrum or in conjunction with other analytical spectra to confirm the structure.