Analysis of Compounds Dissolved in Nonpolar Solvents by Electrospray Ionization on Conductive Nanomaterials
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Electrospray ionization mass spectrometry (ESI-MS) technique has limitations in analysis of compounds that are dissolved in nonpolar solvents. In this study, ambient ionization of compounds in solvents that are not “friendly” to electrospray ionization, such as n-hexane, is achieved by conductive nanomaterials spray ionization (CNMSI) on nanomaterial emitters, including carbon nanotubes paper and mesodendritic silver covered metal, which applies high voltages to emitters made of these materials without the assistance of polar solvents. Although the time intensity curves (TIC) commonly vary from 4.5% to 23.7% over analyses, protonated molecular ions were found to be the most abundant species, demonstrating good reproducibility of the technique in terms of ionized species. Higher mass spectrometric responses are observed in analyzing nonpolar systems than polar systems. 2-Methoxyacetophenone, 4-methylacetophenone, benzothiazole, quinolone, and cycloheptanone as low as 2 pg in n-hexane can be directly detected using the developed method. The developed technique expands the analysis capability of ESI-MS for direct, online analysis of nonpolar systems, such as low polarity extracts, normal phase liquid chromatography eluates, and synthetic mixtures.
KeywordsConductive nanomaterials Electrospray ionization Nonpolar solvents
As a powerful mass spectrometric technique, electrospray ionization (ESI) has been demonstrated to have significant advantages in speed and sensitivity among other chemical analysis methods, and it has been widely applied in many scientific fields to analyze a great variety of compounds. Traditionally, these compounds are dissolved in aqueous mixtures of polar organic solvents (e.g., methanol and acetonitrile) prior to ESI. However, ESI is limited to analyzing compounds dissolved in nonpolar solvents due to the solvents’ poor conductivities and low dielectric constants. As a result, the nonpolar solvents that are widely used in organic reactions, normal phase liquid chromatography (NPLC), gel permeation chromatography (GPC), sample extraction and NMR analysis, are less amenable for use in ESI-MS. To analyze nonpolar systems using ESI-MS, adding polar solvents or ionic liquids to solution was reported to facilitate ionization of dissolved species [1, 2, 3]. However, it changes the solvent system of the samples that may lead to deviations from the prior conditions.
In recent years, much effort has been made, in combination with the emergence of ambient ionization techniques, to detect chemicals in nonpolar solvents. Previous research on recently developed solvent-assisted, electrospray-based ionization methods illustrated that extractive electrospray ionization (EESI) [4, 5], solvent-assisted electrospray ionization (SAESI) , and continuous flow-extractive desorption electrospray ionization (CF-EDESI) [7, 8] could be used for the direct analysis of compounds in “non-ESI-friendly” solvents. In these ionization methods, introduction of the nonpolar system’s sample and ionization are independent processes, which prevent the sample conditions from changing. However, complicated instrument modifications or complex parameter adjustments to these methods are still required. In addition to solvent-assisted ESI, solid substrate ESI techniques such as wooden tip ESI [9, 10, 11], probe electrospray ionization (PESI) [12, 13, 14, 15, 16], and paper spray ionization (PSI) [17, 18, 19] have been extended to directly sample and ionize various types of samples. By first sampling and then subjecting samples to ESI-MS analysis with the assistance of polar solvents or polar solvent vapor would be a promising approach for these techniques to analyze analytes in nonpolar solvents. It is worth noting that among the above methods, paper substrate-based PSI is capable of directly ionizing insoluble drugs, peptides, nucleotides, and phospholipids as solids from paper wetted with nonpolar solvent . Nevertheless, according to our repeated trials, the ionization efficiency of PSI is quite limited concerning such use.
Discovered in 1991 , carbon nanotubes (CNTs) have been recognized for their low weight , good heat conductance , various electronic properties that depend on their structure, and the ability to transfer charge at relatively low voltages due to their high aspect ratios and nanometer-sized tips . Recently, CNTs have mainly been used in matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS) as a matrix . In addition, based on CNTs, field emission, chemical ionization (CI) , electron ionization (EI) , as well as field emission ionization  were developed. In ESI field, paper substrates are capable of generating spray ionization under an electronic potential of just a few volts after the paper is coated with CNTs . We speculate the conductive nano-structure (as that of CNTs) may play an important role in facilitating ionization, and thus may be used in ionization of “non-ESI-friendly” systems. However, the potential of using conductive nano-materials for the ionization of analytes dissolved in nonpolar solvents has not yet been investigated. After evaluating many materials in our preliminary experiments, we ultimately found that ionization efficiency of “non-ESI-friendly” systems significantly improved compared with conventional technologies when using conductive nanomaterials such as CNTs and mesodendritic silver-covered metal as ESI emitters.
In this study, we report the developed use of conductive nanomaterials spray ionization mass spectrometry (CNMSI-MS) in the analysis of analytes in nonpolar solvents. Conductive nanomaterial is first cut into a triangle, and a silica capillary connected to a syringe pump is used to supply analyte solution to the center of the triangle through contact with its surface. Analytes transport to the triangle tip via capillary action through micro-channels in the emitter substrate, and then a high electric field is applied to perform ionization. CNMSI-MS studies of compounds dissolved in solutions, such as methanol, n-hexane, and dichloromethane (CH2Cl2), were investigated. Chiral molecules separated by NPLC were also detected by CNMSI-MS without any polar assistant solvent, and intense signals of [M+H]+ ions from each racemic compound is clearly observed in positive ionization mode.
Perphenazine (1) and 5-acrylamide-1,10-phenanthroline (2) were synthesized in our laboratory and confirmed structurally using UV spectroscopy, MS, 1H NMR, and 13C NMR. The purity of each compound was verified as being higher than 98% using HPLC–UV analysis. 2-Methoxyacetophenone (3), 4-methylacetophenone (4), 1-indanone (5), acetophenone (6), 4'-dimethylaminoacetophenone (7), 5-methylindole (8), benzothiazole (9), quinolone (10), cycloheptanone (11), 2,2'-dipyridyl (12), and silver nitrate (13) were purchased from Aladdin Reagent Co., Ltd. (Shanghai, China). Single-walled carbon nanotubes (SWCNTs) were obtained from Aladdin Reagent Co., Ltd. (Shanghai, China). Grade ET31 chromatography paper (0.50 mm thick) was purchased from Whatman International Ltd. (Maidstone, England). Water was prepared using a Milli-Q system (Millipore Laboratory, Bedford, MA, USA). HPLC-grade methanol, acetonitrile, ethanol, and acetone were obtained from Merck Co. (Darmstadt, Germany). n-Hexane and CH2Cl2 of analytical reagent grade were obtained from Changzheng Chemical Reagent Corp. (Chengdu, China). HPLC grade methanol was purchased from Fisher Co. (Fisher Scientific, Hampton, NH, USA). N-(2-(hydroxymethyl)-1-(2-methoxyphenyl)-3-methylbut-3-en-1-yl)P,P-diphenylphosphinic amide raceme (14) was synthesized in our laboratory. Other reagents were of analytical grade and used without further purification.
Preparation of CNTs Sheet and Mesodendritic Silver Covered Metal
CNTs sheet (10–50 μm thick) was produced by multiple steps of tube dispersion and suspension filtration as described previously . SWCNTs were sonicated for 5 h in 5 M hydrochloric acid (HCl) to dissolve residual iron catalyst particles. Concentrated nitric acid (HNO3) was added to the acid etch and kept between 55 and 60 °C for 5 h to increase the effectiveness of the purification and populate the nanotube surface with carboxylic acid groups. The material was neutralized through a series of deionized water washes, then filtered, dried, and reground into powder. Using nanosperse AQ surfactant (Waltham, MA, USA), the purified nanotubes were suspended in deionized water and sonicated. An aliquot of the suspension was then removed and vacuum-filtered through a microporous membrane (0.45 μm) to fabricate a free-standing, round film of CNTs. Subsequent washes with deionized water and methanol were performed. After washing, the CNTs sheet was carefully peeled off from the filter membrane. Finally, the paper was dried in an oven at 30 °C for 24 h in vacuo to remove excess moisture.
Mesodendritic silver-covered zinc sheet was synthesized in the same procedure as reported in our previous work .
The study was performed on a Waters Xevo TQ mass spectrometer (Waters, Manchester, UK) equipped with an orthogonal Z-spray electrospray ionization source (ESI). For conventional ESI-MS experiments, the parameters were set as follows: 500 L/h desolvation gas flow rate; 500 °C desolvation gas temperature; 150 °C source temperature; 3 kV capillary voltage; and 30 V cone voltage. The accumulation time for the ion signal to acquire a single mass spectrum record was set to 0.2 s. Tandem mass spectra were obtained by collision-induced dissociation (CID) of selected precursor ions with argon as collision gas. For CNMSI-MS experiments, desolvation and nebulization gases were turned off. Because an external high voltage supply was used for ionization, the capillary voltage was also turned off. Other parameters were the same as those for conventional ESI-MS experiments. Data acquisition and processing were performed using Masslynx 4.1 software (Waters Corp., Milford, MA, USA). All experiments were carried out in triplicate or more in an air-conditioned room with constant temperature and humidity.
Results and Discussion
Characterization of CNTs Paper
Evaluation Experiments of Conductive Nanomaterials
A series of evaluation experiments were conducted using CNMSI-MS analysis of standard compounds dissolved in different solutions. The emitter tip-to-MS inlet distance and spray voltage were investigated.
The Practical Application of Low Polarity Compounds Analyzed by CNMSI-MS Methods
CNMSI-MS can ionize chemicals dissolved in nonpolar solvents, such as hexane, without any assistance from polar solvents. The developed ambient ionization technique expands the analytic capability of ESI-MS to the direct, online analysis of nonpolar systems, such as low polarity extracts, NPLC elutes, and synthetic mixtures. Compared with conventional PSI-MS, CNMSI-MS produces superior signals in terms of ion intensity and limit of detection, indicating the higher ionization efficiency and sensitivity of this method for analytes in nonpolar solvents. Field desorption from nano-structures within the conductive nanomaterials is presumed to be one of the key factors. Further applications of this technology are ongoing, and should broaden the analytical scope of a commercial mass spectrometer with only minor modifications.
This research was supported by the National Natural Science Foundation of China (no. 21672206 and 21572221).
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