Pressure Chemical Ionization MS Research Articles

University teaching of mass spectrometry as a key practical technique within the context of a fully integrated, spiral curriculum

RationaleMass spectrometry (MS) is introduced to high school students in the UK in many pre‐university course syllabi. As such, we have identified the use of MS as a key technique that should be taught practically to undergraduates from the outset of their studies. This mini‐review describes how we introduce and develop students' use of MS throughout our three‐year undergraduate spiral curriculum practical programme, using atmospheric pressure chemical ionization MS (APCI‐MS).MethodsWe have used an Advion ExpressionL spectrometer, fitted with an atmospheric solids analysis probe or a Plate Express TLC sampler for sample introduction.ResultsWe have successfully demonstrated the use of APCI‐MS in a range of practicals and experiments covering organic and organometallic chemistries, with large cohorts of students gaining hands‐on instrumental experience in authentic research settings.ConclusionsAPCI‐MS has proven to be an easy‐to‐use and valuable addition to our undergraduate practical course. The robustness of the spectrometer enables routine use by large cohorts of students with minimal supervision, and routine maintenance can be carried out by non‐specialist technicians. Students can readily process and interpret results for a series of routine analyses, as well as demonstrate uses in problem‐solving exercises.

Formation of Previously Undescribed Δ7-Phytosterol Oxidation Products and Tocopherylquinone Adducts in Pumpkin Seed Oil during Roasting, Screw-Pressing, and Simulated Culinary Processing at Elevated Temperatures.

The influence of pumpkin seed roasting conditions (110-140 °C) and screw-pressing on the formation of previously undescribed Δ7-phytosterol oxidation products and tocopherylquinone adducts with nucleophilic phosphatidylethanolamine species was investigated. The roasting process of pumpkin seed paste at a temperature above 120 °C for 30 min considerably enhanced the formation of Δ7-oxysterols. Targeted analysis [electron impact mass spectrometry (MS), 1D-nuclear magnetic resonance] led to the identification of five novel markers of pumpkin paste roasting, among which (3β,5α,22E,24S)-stigmasta-7,22-dien-6-one-3-ol (6-oxo-α-spinasterol), stereoisomers of (3β,5α,22E)-7,8-epoxystigmast-22-en-3-ol (7,8-epoxy-α-spinasterol), and (3β,5α)-22,23-epoxystigmast-7-en-3-ol (7,8-epoxy-α-spinasterol) were reported in edible oils for the first time. Simulated culinary processing provided novel stereoisomers of (3β,5α,22E)-stigmasta-7,22-dien-3,6-diol, unusual (3β,5α,22E)-stigmasta-7,22-dien-6,15-dione-3-ol, and (5α,22E)-stigmasta-7,22-dien-3-one accompanied by minor stereoisomers of (3β,5α)-7,8;22,23-diepoxystigmastan-3-ol. Moreover, a clear relationship between the pumpkin seed oil stability index and synergistic effect of glycerophospholipids with present tocochromanols was found. High-resolution atmospheric pressure chemical ionization-MS experiments clearly demonstrated the formation of various γ-tocopherylquinone adducts with primary amines, namely, octylamine. The mitigation strategy of potentially detrimental oxysterols from pumpkin seed oil included optimization of processing parameters while maintaining the formation of desirable sensory-active compounds.

Establishment of a GC-MS-based 13 C-positional isotopomer approach suitable for investigating metabolic fluxes in plant primary metabolism.

13 C-Metabolic flux analysis (13 C-MFA) has greatly contributed to our understanding of plant metabolic regulation. However, the generation of detailed in vivo flux maps remains a major challenge. Flux investigations based on nuclear magnetic resonance have resolved small networks with high accuracy. Mass spectrometry (MS) approaches have broader potential, but have hitherto been limited in their power to deduce flux information due to lack of atomic level position information. Herein we established a gas chromatography (GC) coupled to MS-based approach that provides 13 C-positional labelling information in glucose, malate and glutamate (Glu). A map of electron impact (EI)-mediated MS fragmentation was created and validated by 13 C-positionally labelled references via GC-EI-MS and GC-atmospheric pressure chemical ionization-MS technologies. The power of the approach was revealed by analysing previous 13 C-MFA data from leaves and guard cells, and 13 C-HCO3 labelling of guard cells harvested in the dark and after the dark-to-light transition. We demonstrated that the approach is applicable to established GC-EI-MS-based 13 C-MFA without the need for experimental adjustment, but will benefit in the future from paired analyses by the two GC-MS platforms. We identified specific glucose carbon atoms that are preferentially labelled by photosynthesis and gluconeogenesis, and provide an approach to investigate the phosphoenolpyruvate carboxylase (PEPc)-derived 13 C-incorporation into malate and Glu. Our results suggest that gluconeogenesis and the PEPc-mediated CO2 assimilation into malate are activated in a light-independent manner in guard cells. We further highlight that the fluxes from glycolysis and PEPc toward Glu are restricted by the mitochondrial thioredoxin system in illuminated leaves.

Simultaneous analysis of 21 bioactive compounds in biorefinery oil: Multivariate optimization of a method based on liquid chromatography, atmospheric pressure chemical ionization and tandem mass spectrometry

A HPLC method coupled to atmospheric pressure chemical ionization (APCI) and tandem MS was optimized for the analysis of tocotrienols, tocopherols, retinol, squalene, phytosterols and carotenoids in lipid matrices. The aim of the research was to obtain a specific, rapid and robust method for bioactive compounds in by-products from the agri-food industry, in order to discover new high added-value products.The separation of 21 analytes, by non-aqueous reverse phase (NARP) liquid chromatography, was optimized by using design of experiments and response surface methodology. The best compromise among peak resolution and rapidity consisted in a run of 25 min. The problem of coelution of some substances was overcome thanks to the mass detection in multiple reaction monitoring mode, which also allowed maximization of sensitivity. The instrumental method was characterized by calibration curves with R2 of 0.989–0.999, limits of detection in the range 0.6–27 µg L-1, relative standard deviation always <20% and excellent specificity (4 identification points for each compound). The method was applied, with minimal sample pre-treatment, to vegetable oil mixtures and processed products from a first-generation biodiesel plant. Negligible matrix effect and accuracy of 80–90% were observed for most analytes, thanks to the use of APCI and dilution factors ranging from 500 to 7500 (volume to mass ratio). The most concentrated species were phytosterols, squalene and tocopherols, which showed levels of 10–30 mg g−1 in some samples, indicating the possibility of exploiting these by-products for the recovery of high-value chemicals.

Pagoda[5]arene with Large and Rigid Cavity for the Formation of 1∶2 Host–Guest Complexes and Acid/Base-Responsive Crystalline Vapochromic Properties

Pagoda[5]arene (P5), which is composed of five 2,6-dimethoxylanthracene (2,6-DMA) subunits bridged by methylene groups at 1,5 positions, was conveniently synthesized in 43% by trifluoroacetic acid ...

Metabolite Profiling of Manilkara zapota L. Leaves by High-Resolution Mass Spectrometry Coupled with ESI and APCI and In Vitro Antioxidant Activity, α-Glucosidase, and Elastase Inhibition Assays.

High-resolution mass spectrometry equipped with electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) sources was used to enhance the characterization of phytochemicals of ethanol extracts of Manilkara zapota L. leaves (ZLE). Sugar compounds, dicarboxylic acids, compounds of phenolic acids and flavonoids groups, and other phytochemicals were detected from the leaves. Antioxidant activity and inhibition potentiality of ZLE against α-glucosidase enzyme, and elastase enzyme activities were evaluated in in vitro analysis. ZLE significantly inhibited activities of α-glucosidase enzyme at a lower concentration (IC50 2.51 ± 0.15 µg/mL). Glucose uptake in C2C12 cells was significantly enhanced by 42.13 ± 0.15% following the treatment with ZLE at 30 µg/mL. It also exhibited potential antioxidant activities and elastase enzyme inhibition activity (IC50 27.51 ± 1.70 µg/mL). Atmospheric pressure chemical ionization mass spectrometry (APCI–MS) detected more m/z peaks than electrospray ionization mass spectrometry (ESI–MS), and both ionization techniques illustrated the biological activities of the detected compounds more thoroughly compared to single-mode analysis. Our findings suggest that APCI along with ESI is a potential ionization technique for metabolite profiling, and ZLE has the potential in managing diabetes by inhibiting α-glucosidase activity and enhancing glucose uptake.

Formation of the exceptional [M - H]+ cation in atmospheric pressure ionization mass spectrometry analysis of 2-(diphenylsilyl) cyclopropanecarboxylate esters.

In general, ionization of analytes in atmospheric pressure ionization mass spectrometry (API-MS) in positive ion mode results in the formation of protonated molecules ([M + H]+ ) and/or cationized molecules (e.g., [M + Na]+ ). The formation of specific [M - H]+ cations in the API process is of significant interest for further investigation. The ionization processes of 2-(diphenylsilyl)-1-phenyl-cyclopropanecarboxylate esters were investigated using electrospray ionization (ESI)-MS and atmospheric pressure chemical ionization-MS in positive ion mode. Theoretical calculations were carried out with the Gaussian 03 program using the density functional theory (DFT) method at the B3LYP/6-311 + G(2d,p) level. The anomalous [M - H]+ ion and the regular [M + Na]+ ion were both observed using ESI-MS. Interestingly, no [M + H]+ ion was obtained in the ESI-MS analysis, and acidification of the ESI solvent accelerated the formation of [M - H]+ rather than [M + H]+ ion. DFT calculations for the typical methyl 2-(diphenylsilyl)-1-phenyl-cyclopropanecarboxylate (1) indicated that the [1 + H]+ ion can thermodynamically and kinetically undergo facile H2 elimination to generate [1 - H]+ . The favorable formation of [M - H]+ ions in these compounds is attributed to the unique diphenylhydrosilyl group in their structure. The [M + H]+ ion formed easily underwent H2 elimination to produce the [1 - H]+ ion in the API source, and thus, acidification of the ESI solvent apparently accelerates the formation of the [1 - H]+ ion.

Regioisomeric and enantiomeric analysis of primary triglycerides in human milk by silver ion and chiral HPLC atmospheric pressure chemical ionization-MS

Triglycerides (TG) not only provide energy for infants but have important physiological functions. Understanding the composition and structure of TG in human milk is conducive to the development of infant formulas. In this study, TG species in human milk from 3 provincial capitals (Zhengzhou, Wuhan, and Harbin) in different regions of China were determined through C18 HPLC electrospray ionization tandem mass spectrometry (MS). The results showed that in human milk from these 3 regions, oleoyl-palmitoyl-linoleoylglycerol (OPL; 16.55, 19.20, and 18.67%, respectively) was more abundant than oleoyl-palmitoyl-oleoylglycerol (OPO; 10.08, 10.22, and 12.03%, respectively). Subsequently, regioisomeric and enantiomeric analysis of main TG in the human milk were performed on silver ion and chiral HPLC atmospheric pressure chemical ionization mass spectrometry (APCI)-MS, respectively. The results showed that rac-OPL (above 85%), rac-OPO (above 85%), rac-palmitoyl-oleoyl-oleoylglycerol (PPO; above 90%), and rac-OLaO (above 70%) were the main regioisomers of OPL, OPO, PPO, and lauroyl-oleoyl-oleoylglycerol (LaOO), respectively. The relative ratios of enantiomer pairs of rac-OPL (rac-OPL1 and rac-OPL2) were about 37 and 63%, respectively.

Development of a liquid chromatography atmospheric pressure chemical ionization mass spectrometry method for determining off-flavor compounds and its application toward marine recirculating aquaculture system monitoring and evaluation of aeration as a depuration approach

The off-flavor compounds geosmin and 2-methylisoborneol (2-MIB) are well-known to impact the quality of farmed freshwater fish species, but little is known about off-flavors in marine aquaculture. To begin addressing this knowledge gap, a method for determining geosmin and 2-MIB using LC with atmospheric pressure chemical ionization (APCI) MS detection was developed. While 2-MIB was readily detected using LC-APCI/MS, geosmin exhibited on-column degradation that was independent of column chemistry and could not be eliminated. Optimized conditions were identified that balanced the separation and ionization efficiency of 2-MIB and geosmin while minimizing geosmin degradation, but the overall method sensitivity for geosmin was reduced by the on-column losses. The method was used with direct aqueous injections to determine the volatilization rates of geosmin and 2-MIB at ppb levels during aeration under laboratory conditions in both salt water and pure water to simulate marine and fresh water aquaculture, respectively. The volatilization rates of both compounds were 30% faster in salt water than in fresh water with or without aeration, but aeration was found to enhance the rate by a factor of 2.5 in both water types. The LC-APCI/MS method was combined with stir bar sorptive extraction (SBSE) to achieve greater sensitivity for determining off-flavors in recirculating aquaculture system (RAS) water. Using SBSE-LC-APCI/MS, the LODs for geosmin and 2-MIB were 70 ng/kg (part per trillion) and 6 ng/kg, respectively. The on-column losses resulted in a relatively high LOD for geosmin that renders this method unsuitable for determining geosmin at the low ng/kg levels expected in RAS. SBSE using both grab water samples and an in-situ diving unit were used to evaluate 2-MIB levels in the culture water of two separate marine RAS that were supporting the growth of European sea bass but had differing levels of water treatment. 2-MIB was readily detected using both SBSE approaches in the RAS with less sophisticated treatment when the animal stocking density was at its highest (50 kg/m3) but was not detected in the more sophisticated RAS regardless of stocking density. Geosmin was not detected in either system, but the results were inconclusive given its higher LOD. These limited results suggest that the anaerobic water treatment components, present only in the more sophisticated RAS, maintained the level of 2-MIB below the LOD.

Detection of Edible Plant Oil Adulteration by Triacylglycerol Profiles Using an Atmospheric Pressure Chemical Ionization Source and MS3 Ion Trap Mass Spectrometry

An adulteration of high‐price oil has been an important concern in agri‐food fraud problems. Triacylglycerols are the main components of edible plant oils, which play an important role as target for adulteration detections. The objective is to evaluate the utility of triacylglycerol profiles in the detection of adulteration in high priced oils. An effective method is established to detect adulteration of high priced oils based on triacylglycerol profiles and chemometrics. Triacylglycerol profiles of edible oils are analyzed by an APCI‐MS3‐IT‐MS. All triacylglycerol compounds are quantified by normalization of the chromatographic peak area in selected reaction monitoring (SRM) mode based on MS3 fragment ion pairs, which are used to eliminate interference from the isobaric species of triacylglycerol. The results of the hierarchical cluster analysis (HCA) and random forest (RF) classification algorithm indicated that peanut, soybean, sesame, sunflower seed, and linseed oils are completely classified into five groups based on their triacylglycerol profiles. Finally, a recursive support vector machine (R‐SVM) discriminant model is established, which can successfully identify adulteration of high priced oil with cheaper edible oils at a concentration of as low as 4% with an accuracy of 93.7%.Practical Applications: The triacylglycerol profiles of edible plant oils are analyzed by RPLC‐APCI‐IT‐MS. The SRM scan mode based on MS3 fragment ion pairs can eliminate interference of triacylglycerol isomers. The discriminant model for adulterated high‐price oils is established by R‐SVM. HCA, and RF can classify the five kinds of edible plant oil.The triacylglycerol of five edible plant oils is analyzed by RPLC‐APCI‐MS3‐IT‐MS. The triacylglycerol data matrix is submitted using unsupervised (HCA) and supervised (RF) chemometric methods to classify five kinds of edible plant oil. A discriminant model established with the R‐SVM can be used to identify adulterated linseed oil samples (≥4%) with an accuracy of 93.7%.

Fatty Acid, Triacylglycerol and Unsaponifiable Matters Profiles and Physicochemical Properties of Chinese Evening Primrose Oil.

This work focused on physicochemical property assaying, fatty acid composition, triacylglycerol (TAG) profiles, and unsaponifiable matters composition of the Chinese evening primrose oil. The cold press oil possessed very low acid value and peroxide value, and relatively high iodine value. Fatty acid composition results indicated that this oil was especially high in linoleic acid and linolenic acid. Characterization of TAG composition was achieved by a two-dimensional HPLC coupling of nonaqueous reverse-phase and silver ion HPLC with atmospheric pressure chemical ionization MS method. There was a total of 38 TAGs including 27 regioisomers which had been determined. Unsaponifiable matters composition results revealed that this oil possessed a number of phytosterols, in which β-sitosterol and stigmasterol were most predominant.

Simultaneous Quantification of Free Cholesterol, Cholesteryl Esters, and Triglycerides without Ester Hydrolysis by UHPLC Separation and In-Source Collision Induced Dissociation Coupled MS/MS

We demonstrate the application of in-source nitrogen collision-induced dissociation (CID) that eliminates the need for ester hydrolysis before simultaneous analysis of esterified cholesterol (EC) and triglycerides (TG) along with free cholesterol (FC) from human serum, using normal phase liquid chromatography (LC) coupled to atmospheric pressure chemical ionization (APCI) tandem mass spectrometry (MS/MS). The analysis requires only 50 μL of 1:100 dilute serum with a high-throughput, precipitation/evaporation/extraction protocol in one pot. Known representative mixtures of EC and TG species were used as calibrators with stable isotope labeled analogs as internal standards. The APCI MS source was operated with nitrogen source gas. Reproducible in-source CID was achieved with the use of optimal cone voltage (declustering potential), generating FC, EC, and TG lipid class-specific precursor fragment ions for multiple reaction monitoring (MRM). Using a representative mixture of purified FC, CE, and TG species as calibrators, the method accuracy was assessed with analysis of five inter-laboratory standardization materials, showing –10% bias for Total-C and –3% for Total-TG. Repeated duplicate analysis of a quality control pool showed intra-day and inter-day variation of 5% and 5.8% for FC, 5.2% and 8.5% for Total-C, and 4.1% and 7.7% for Total-TG. The applicability of the method was demonstrated on 32 serum samples and corresponding lipoprotein sub-fractions collected from normolipidemic, hypercholesterolemic, hypertriglyceridemic, and hyperlipidemic donors. The results show that in-source CID coupled with isotope dilution UHPLC-MS/MS is a viable high precision approach for translational research studies where samples are substantially diluted or the amounts of archived samples are limited.Graphical ᅟ

Molecularly imprinted solid-phase extraction coupled with LC-APCI-MS-MS for the selective determination of serum cholesterol.

The assessment of selective molecularly imprinted solid-phase extraction (MISPE) for clean-up and extraction of cholesterol (CHO) from bovine and human serum samples was evaluated. Two molecular imprinted polymers (MIP) based on organic and inorganic/organic (hybrid) materials were obtained and compared regarding recovery and selectivity toward CHO. Optimum chromatographic separation between the CHO and its isobaric compound lathosterol (LATHO) has been achieved within 15min using an Agilent Poroshell 120 EC-C18 column (120mm×3.0mm, 2.7μM) and mobile phase consisting of acetonitrile:methanol (8:2, v/v) at a flow rate of 0.6mLmin-1 . Mass spectrometry detection was operated in atmospheric pressure chemical ionization (APCI) (+) mode and the SIM transitions [M-H2 O+H]+ for CHO and LATHO were 364.1 → 161.2 → 95.2 and 364.1 → 95.1 → 81.1m/z, respectively. The sample preparation could be accomplished within 1h, and the matrix effect on the MS signal after sample pretreatment did not interfere in the analyses. The method provided a good linear range (0.013-312μM.) with coefficients of determination of R2 >0.99, substantial sensitivity (limits of detection of 6.5nM), good recovery (∼90%), high reproducibility (intraday and interday relative standard deviations less than 15%) and satisfactory accuracy (relative error ranged from -13.46% to 11.54%). The organic MIP showed superior performance compared with the hybrid. Finally, the MISPE procedure followed by LC-APCI-MS-MS analysis showed their suitable applicability to examine CHO and LATHO in serum samples.

Lipidomic analysis of plasma, erythrocytes and lipoprotein fractions of cardiovascular disease patients using UHPLC/MS, MALDI-MS and multivariate data analysis.

Differences among lipidomic profiles of healthy volunteers, obese people and three groups of cardiovascular disease (CVD) patients are investigated with the goal to differentiate individual groups based on the multivariate data analysis (MDA) of lipidomic data from plasma, erythrocytes and lipoprotein fractions of more than 50 subjects. Hydrophilic interaction liquid chromatography on ultrahigh-performance liquid chromatography (HILIC-UHPLC) column coupled with electrospray ionization mass spectrometry (ESI-MS) is used for the quantitation of four classes of polar lipids (phosphatidylethanolamines, phosphatidylcholines, sphingomyelins and lysophosphatidylcholines), normal-phase UHPLC-atmospheric pressure chemical ionization MS (NP-UHPLC/APCI-MS) is applied for the quantitation of five classes of nonpolar lipids (cholesteryl esters, triacylglycerols, sterols, 1,3-diacylglycerols and 1,2-diacylglycerols) and the potential of matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) is tested for the fast screening of all lipids without a chromatographic separation. Obtained results are processed by unsupervised (principal component analysis) and supervised (orthogonal partial least squares) MDA approaches to highlight the largest differences among individual groups and to identify lipid molecules with the highest impact on the group differentiation.

Identification of hydrolytic and isomeric N-oxide degradants of vilazodone by on line LC–ESI–MS/MS and APCI–MS

The present study reports the degradation behavior of a new antidepressant drug, vilazodone, under various stress conditions as per International Conference on Harmonization guidelines (ICH, Q1A(R2). The investigation involved monitoring decomposition of the drug under hydrolytic (acidic, basic and neutral), oxidative, photolytic and thermal stress conditions and identifying degradation products. A rapid, precise, accurate and robust ultra high performance liquid chromatography (UPLC) method has been developed on a Waters CSH Phenyl-Hexyl column (100mm×2.1mm, 1.7μm) using gradient elution of 10mM ammonium acetate buffer (pH 5.0) and acetonitrile as mobile phase. The drug was found to be degraded in hydrolytic (acidic and basic) and oxidative conditions, whereas it was stable under neutral hydrolytic, photolytic and thermal stress conditions. The method was extended to quadrupole time-of-flight mass spectrometry (QTOF-MS) for the structural characterization of degradation products. It has been observed that isomeric N-oxide degradation products were formed under oxidative stress condition. The exact location of N-oxidation in the drug was investigated using atmospheric pressure chemical ionization (APCI) due to the formation of characteristic fragment ions. These fragment ions resulted from Meisenheimer rearrangement owing to thermal energy activation at the vaporizer of APCI source. All degradation products were comprehensively characterized by UPLC–ESI–MS/MS and UPLC–APCI–MS experiments. The most probable mechanisms for the formation of degradation products have also been proposed. The method was validated in terms of specificity, linearity, accuracy, precision, and robustness as per ICH guidelines.

Identification of regioisomers and enantiomers of triacylglycerols in different yeasts using reversed‐ and chiral‐phase LC–MS

LC with atmospheric pressure chemical ionization (ACPI) MS with RP and chiral phase was used for separation of triacylglycerols (TAGs) from yeasts of the genera Candida, Kluyveromyces, Rhodotorula, Saccharomyces, Torulospora, Trichosporon, and Yarrowia. Chiral LC-APCI-MS is based on using two columns in series packed with a 3,5-dimethylphenyl carbamate modified β-cyclodextrin chiral phase. All regioisomers and enantiomers of TAGs containing one to five double bonds were separated. Molecular species of TAGs, i.e. regioisomers and enantiomers, were identified and quantified by MS/MS. Among the 94 identified TAGs, the most abundant were triolein, oleopalmitoleoolein, and dipalmitoleoolein. In strains producing palmitoleic acid in amounts >25% of total fatty acids (FAs), this acid, or unsaturated FA is bound in sn-1. In strains containing palmitoleic acid at 10-25% total FAs this acid is mainly bound in sn-3, saturated FA being bound in sn-1. Strains containing <10% palmitoleic acid form preferentially symmetrical TAGs.

A Rapid and Sensitive LC–MS Method for Determination of Ezetimibe Concentration in Human Plasma: Application to a Bioequivalence Study

A selective and highly sensitive high performance liquid chromatography-electrospray ionization mass spectrometry method has been developed for determination of ezetimibe concentrations in human plasma. Ezetimibe was extracted from plasma with ethyl acetate followed by evaporation of the organic layer and, then, reconstitution of the residue in mobile phase before injection to chromatograph. The mobile phase consisted of acetonitrile-ammonium acetate (10 mM, pH 3.0), 75:25 (v/v). An aliquot of 10 μL was chromatographically analyzed on a prepacked Zorbax XDB-ODS C18 column (2.1 × 100 mm, 3.5 micron). Detection of analytes was achieved by mass spectrometry with atmospheric pressure chemical ionization (APCI) interface in the negative ion mode operated under the multiple-reaction monitoring mode (m/z transition: ezetimibe 408–271). Standard curves were linear (r = 0.998) over the wide ezetimibe concentration range of 0.05–30.0 ng mL−1 with acceptable accuracy and precision. The limit of detection was 0.02 ng mL−1. The validated LC–APCI–MS method has been used successfully throughout a bioequivalence study on an ezetimibe generic product in 24 healthy male volunteers.

Characterization and quantification of triacylglycerols in peanut oil by off‐line comprehensive two‐dimensional liquid chromatography coupled with atmospheric pressure chemical ionization mass spectrometry

The complexity of natural triacylglycerols (TAGs) in various edible oils is prodigious due to the hundreds of set is of TAG compositions, which makes the identification of TAGs quite difficult. In this investigation, the off-line 2D system coupling of nonaqueous RP and silver-ion HPLC with atmospheric pressure chemical ionization MS detection has been applied to the identification and quantification of TAGs in peanut oil. The method was successful in the separation of a high number of TAG solutes, and the TAG structures were evaluated by analyzing their atmospheric pressure chemical ionization mass spectra information. HPLC and MS conditions have been optimized and the fragmentation mechanisms of isomers have been validated. In addition, an internal standard approach has been developed for TAG quantification. Then this system was applied in peanut oil samples and there was a total of 48 TAGs including regioisomers that have been determined and quantified.

Normal phase LC coupled with direct analysis in real time MS for the chiral analysis of 4‐(methylnitrosamino)‐1‐(3‐pyridyl)‐1‐butanol and jasmonic acid

Normal phase chiral LC (NPLC) has been proved to be powerful and efficient for chiral separation. However, the combination of NPLC with ESI or atmospheric pressure chemical ionization MS is restricted by the poor ionization efficiency and thermal fragmentations of analytes to some extent. Direct analysis in real time MS (DART-MS) is an ambient ionization technique that shows high ionization efficiency of the analytes in the normal phase mobile phase. In this work, we coupled chiral NPLC to DART-MS for the chiral qualitative and quantitative analysis of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol and jasmonic acid enantiomers. Satisfactory results for the enantiomers of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol operating in the positive mode were obtained in terms of linearity (2.5-250 μg/mL, R(2) , 0.999-1.000) and repeatability (25 μg/mL, RSDs, 4.7-5.6%). Moreover, chiral NPLC-DART-MS resulted in the simultaneous chiral separation and detection of jasmonic acid enantiomers, which are very difficult to be analyzed by NPLC-ESI-MS and NPLC-APCI-MS. Compared with the coupled techniques of NPLC-ESI-MS and NPLC-APCI-MS, NPLC-DART-MS showed advantages in increasing the ionization efficiency and reducing the in-source thermal fragmentation of analytes.

Regiospecific analysis of TAGs using chromatography, MS, and chromatography‐MS

AbstractIn last two decades, rapid development of analytical methods and their combinations have enabled powerful chromatographical resolution of the regioisomers of TAGs, and their reliable determination by MS using soft ionization methods. In this review article, the resolution of TAG regioisomers by chromatography alone using silver‐ion HPLC, reversed‐phase (RP)‐HPLC, and high‐resolution gas–liquid chromatography (GLC) will be presented. Further, the determination of regioisomers by MS solely, using negative ion chemical ionization tandem MS was reported. In addition, the determination of regioisomers by chromatography and MS using GLC and electron ionization (EI) MS, silver ion HPLC, and atmospheric pressure chemical ionization (APCI) or ESI MS, RP‐HPLC APCI‐MS, and RP‐ and normal‐phase (NP)‐HPLC and ESI‐MS‐MS, are discussed. The use of several LC columns in series, or use of two‐dimensional chromatography has resulted in excellent resolution of the regioisomers of TAGs. The resolution of regioisomers prior to mass spectrometric determination is an essential analytic step, especially, when the composition and structure of short‐chain TAGs, which contain abundantly other isobaric TAG, are analyzed.