A New Matrix For Small Molecule Detection By MALDI-TOF MS Based On Poly-n-vinylcarbazole

Matrix-assisted laser desorption ionisation(MALDI)is a soft ionisation technique developed in the 1980 s.A combination of MALDI with time-of-flight mass spectrometry(TOF MS)is known to rapidly detect nonvolatile,highly polar,and thermally labile molecules,particularly biomacromolecules.Compared with alternative mass spectrometry techniques,MALDI–TOF MS presents numerous advantages,including a high sensitivity,salt tolerance,high throughput,and facile sample preparation.This technique is currently employed for the detection of biomacromolecules such as peptides,proteins,and nucleic acids.In recent years,MALDI–TOF MS has gained considerable attention for small-molecule compound(< 700 Da)analysis applications.However,this technique still faces certain challenges.Among these,one challenge is the difficulty in target spectrum elucidation owing to the severe matrix ion interference of traditional commercialised organic small-molecule matrices in the low-mass region.Another challenge is the uneven crystallisation between traditional organic matrices and analytes,which results in poor repeatability and obstructs efficient quantitative analyses.Thus,it is important to address the above mentioned issues regarding qualitative and quantitative analyses based on MALDI–TOF MS.In this study,the detection of small-molecule compounds via MALDI–TOF MS was improved through the development of a new polymer matrix based on amorphous poly-n-vinylcarbazole(PVK).The experimental details are presented below.1.Amorphous PVK-matrix-based MALDI–TOF MS technique for small-molecule compound detection: herein,the amorphous nitrogen heterocyclic polymer PVK was investigated as a new matrix in MALDI–TOF MS.The analytical performance of PVK for a variety of common small-molecule compounds,including amino acids,nucleic acid bases,fatty acids,environmental endocrine disruptors,and antibiotics,was examined in a negative ion mode.The results revealed that amorphous PVK exhibited wide applicability,high sensitivity,good salt tolerance,and stability for MALDI-based small-molecule compound detection.The detection limits for various small-molecule compounds ranged from0.01–0.0001 mg/m L.The effects of the structure and composition of PVK on the ionisation efficiency of small molecules were explored.The results obtained provide new ideas for the development of related amorphous polymer matrices.2.Amorphous PVK-matrix-based MALDI–TOF MS technique for quantitative detection of enrofloxacin: the performance of the PVK matrix in terms of a quantitative analysis of small-molecule compounds was investigated to establish a fast,high-throughput quantitative MALDI–TOF MS-based analysis technique for enrofloxacin detection in milk samples.This was accomplished based on an isotopic internal standard enrofloxacin-d5 correction method utlizing amorphous polymer PVK as a novel matrix and a representative of enrofloxacin.The method demonstrated good linearity(R2 = 0.9905)in the concentration range of 0.003125–0.8mg/m L.The relative standard deviations were less than 5% for the positive milk samples spiked at concentrations of 0.1 mg/ml.The recoveries in the concentration range of 0.005–0.1mg/m L ranged from 101.5% to 114%,with a detection limit of 0.001 mg/m L and a quantification limit of 0.005 mg/m L.These results indicate that the proposed method is both precise and accurate.3.PVK@SLGO-based MALDI–TOF MS technique for the detection of small-molecule compounds: amorphous PVK was combined with inorganic nanomaterial monolayer graphene oxide(SLGO)to prepare the composite PVK@SLGO.A variety of small-molecule compounds,such as amino acids,nucleic acid bases,environmental endocrine disruptors,and antibiotics,were used to examine the performance of the PVK@SLGO composites as matrices in the positive and negative dual-ion modes for MALDI–TOF MS-based detection of small-molecule compounds.In comparison to PVK,SLGO,and the traditional commercial matrices: 9-AA and CHCA,PVK@SLGO matrices demonstrated good performance and suitability for both positive and negative ion mode detection.PVK@SLGO matrices eliminated interference from their own background peaks,thus enhancing the signal intensity and improving the detection sensitivity.Furthermore,the composite PVK@SLGO matrix exhibited high salt tolerance and stability.The limits of detection for the various small-molecule compounds mentioned above ranged from 0.1–0.0001 mg/m L in the positive ion mode and 0.01–0.0001 mg/m L in the negative ion mode.The structure,composition,and ionisation efficiency of PVK@SLGO were also investigated,thereby laying a foundation for the construction of a dual-mode complex matrix.4.PVK@SLGO-based MALDI–TOF MS technique for the quantification of enrofloxacin:a modified MALDI–TOF MS-based analytical method was used for the quantitative analysis of enrofloxacin in milk samples.This method was based on the isotope internal standard enrofloxacin-d5 correction method in the negative ion mode and utilised the PVK@SLGO composite material as a matrix and enrofloxacin as a representative.The established method demonstrated a good linear relationship(R2 = 0.9991)in the concentration range of 0.0001–1.0mg/m L.The relative standard deviation was within 6% across groups of positive milk samples spiked at a concentration of 0.1 mg/ml.The recovery rate was 100.7%–113.6% within a concentration range of 0.005–0.1 mg/m L.The detection limit of this method was 0.0001mg/m L,and the quantitative limit was 0.005 mg/m L.In summary,this study addressed the current inadequacy of MALDI–TOF MS in detecting the presence of small-molecule compounds by using the amorphous polymer PVK,which served as a new matrix for MALDI-based detection.Consequently,a new method suitable for MALDI-based small-molecule compound detection was constructed.Based on the results,the PVK@SLGO composites were found to be suitable for MALDI positive and negative dual-ion mode detection;moreover,these composites addressed the interference problem of the traditional matrix in the low-mass region.Studies in related areas are highly important to further explore novel matrices and broaden the application scope of MALDI–TOF MS detection techniques.