Quadrupole - Application of Electrostatic Field Orbitrap Mass Spectrometer Q-Exactive in Trace Impurity Inspection

Quadrupole - Application of Electrostatic Field Orbitrap Mass Spectrometer Q-Exactive in Trace Impurity Inspection

- MassFrontier "fish" function to quickly screen and identify related impurities in thiamethoxam

Liu Tian

Thermo Fisher Scientific (China) Co., Ltd.

1. Foreword

Pesticide impurity inspection is an important part of quality control of pesticide products. The identification and monitoring of harmful impurities is of great significance to environmental safety and human health. Q Exactive combines a high-performance quadrupole with high-resolution Orbitrap technology to deliver accurate first-order mass spectral information and high-quality secondary mass spectra. With Mass Frontier 7.0 software for mass spectrometry fragmentation and Fragment Ion Search (FISh) function, the rapid screening and identification of pesticide-related substances has been realized.

Figure 1. Main component thiamethoxam structure

2. Experimental methods

2.1 Instrument:

Mass Spectrometer: Q Exactive Mass Spectrometer

Liquid Chromatograph: Ultimate 3000 UHPLC System

2.2 Test parameters:

Column: Hyper Gold C18 (100 x 2.1mm, 3μm)

Column temperature: 30 ° C

Mobile phase: methanol / 0.1% formic acid water

Flow rate: 0.3 mL/min

Injection volume: 1 μL

gradient:

Ion source: HESI-II,

Source polarity: positive and negative switching

Spray voltage: 3.5KV (+) / 3.2KV (-);

Sheath pressure (N2): 35 arb;

Auxiliary pressure (N2): 10 arb;

Capillary column temperature: 350 ° C;

Gasification temperature: 300 ° C;

Scan mode: one-level full scan (Full Scan, m/z 100~1000) and data-dependent secondary mass spectrometry scan (ddms2);

Resolution: 70,000 (primary mass spectrometry) & 17,500 (secondary mass spectrometry)

Collision mode: high energy collision dissociation (HCD)

Collision energy: NCE30, Stepped NCE50%

2.3 Sample preparation:

Accurately weigh 1mg of thiamethoxam bulk drug, dissolved in methanol to obtain 1mg/mL stock solution. The stock solution was diluted 100 times with methanol as an analytical sample.

3. Results and discussion

3.1 Q-Exactive Quadrupole - The application of the electrostatic field orbitrap mass spectrometer for the inspection of related substances

3.1.1 Positive and negative switching mode

Q-Exactive truly realizes fast switching scan of high resolution mass spectrometry positive and negative ion mode, which effectively improves the measurement efficiency of unknown compounds.

Figure 2. Chromatogram and total ion chromatogram of the PDA obtained from the thiamethoxam sample in positive and negative switching mode (after blank background subtraction)

3.1.2 High quality accuracy

Q-Exactive maintains consistently good mass accuracy during sample determination, with positive and negative ion mode mass deviations less than 1 ppm (external standard method, uncalibrated during test sample) to help you quickly identify unknown material quality information.

Figure 3. Primary component first-order mass spectrum in positive and negative switching mode

3.1.3 High quality MS ² map

HCD (High Energy Collision) provides rich fragment ion information and improves material characterization efficiency.

Figure 4. Secondary mass spectrum in principal component positive ion mode

3.1.4 Wide dynamic range

Q-Exactive accurately measures the mass of different abundance ions with a dynamic range greater than 10e3, ensuring accurate detection of low response components.

Figure 5. Comparison of exact masses for different abundance ions

3.2 thiamethoxam related substance inspection

The primary component secondary mass spectrum was measured by testing the thiamethoxam drug substance sample in positive and negative switching mode (Fig. 4). Mass Frontier 7.0 software can directly import the target compound structure, and after completing the selection of the parent ion ionization mode and the fracture principle, (Fig. 6), the speculation of the mass spectrometry fragment generation of the compound can be quickly completed. (Figure 7).

Mass Frontier 7.0 software can create a compound “Spectral Tree” by directly importing the liquid test raw data file (.raw) and complete the automatic assignment of the target compound's secondary mass spectrometry fragments (Fig. 8). Achieve rapid analysis of compound mass spectrometry fracture rules.

Figure 6. Mass Frontier 7.0 structure editing interface

Figure 7. Mass Frontier 7.0 Compound Mass Fragment Estimation Interface

Figure 8. Mass Frontier 7.0 automates the secondary fragmentation of the target compound

After the analysis of the main component thiamethoxam mass spectrometry, the characteristic fragments of the main component were introduced into the FISh module (Fig. 9) to quickly search for impurities related to the thiamethoxam structure (Fig. 10).

Figure 9. Mass Frontier 7.0 import feature fragment for FISh processing

Figure 10. Mass Frontier 7.0 FISh processing results

The impurity structure identification process is illustrated by taking No. 1 impurity as an example.

Figure 11. Extracted ion flow diagram, primary mass spectrum, and secondary mass spectrum of related substances

From the first-order mass spectrometry information, the measured value of the quasi-molecular ion peak of the related substance No. 1 in the positive ion mode is m/z 248.02548, and the elemental composition is estimated to be C8H11O2N3ClS+, which is different from the main component by the molecular formula N2O. The secondary mass spectrum is analyzed, and the related substance No. 1 has a plurality of fragment ions identical to the main component, indicating that the structure is similar to the main component. In the secondary mass spectrometry cleavage process, thiamethoxam will exhibit characteristic NO2 neutral loss, while No. 1 related substance has no such fragments. Therefore, it is estimated that the structure of related substance No. 1 is shown in Fig. 12.

Figure 12. Structure of the relevant substance No. 1

Using the same method, nine major impurities were detected in the test samples. See Table 1 for details.

Table 1. Main substances related to thiamethoxam

4 Conclusion

Q Exactive was tested on thiamethoxam bulk drug and quickly identified by Mass Frontier 7.0 software to identify 9 major substances, demonstrating Q Exactive's strong application advantages in impurity identification.

Q Exactive uses the Orbitrap quality analyzer to provide ultra-high resolution and good quality accuracy. The hyperbolic quadrupole mass filter improves parent ion selectivity and ion transport efficiency, providing high quality, highly repeatable secondary mass spectrometry. Q Exactive truly achieves fast switching between high-resolution mass spectrometry positive and negative ion modes, which effectively improves the efficiency of qualitative testing.

Q Exactive's good scanning speed is fully adapted to the requirements of UHPLC. This test uses positive and negative switching mode, the first full scan channel uses 70,000 resolution, and the test quality deviation is less than 1ppm (external standard method).

Mass Frontier 7.0 provides great convenience for high-resolution multi-level data processing. On the one hand, it can reasonably speculate the possible fragment ions of the compound and its production pathway through a rich library of mass spectrometry fracture mechanisms; on the other hand, through powerful data processing functions and FISh (feature fragments) Search) function, which quickly filters out useful information from a large number of mass spectral information, finds structurally related compounds, and completes its structural identification.