Regiospecific analysis of diricinoleoylacylglycerols in castor (Ricinus communis L.) oil by electrospray ionization-mass spectrometry

HPLC fractions of diricinoleoylacylglycerols containing one non-ricinoleoyl chain from castor oil were used to identify the regiospecific location of this non-ricinoleoyl chain on the glycerol backbone using electrospray ionization-MS3 of lithium adducts. The regiospecific ions used were from the loss of alpha,beta-unsaturated fatty acid specific at the sn-2 position. The content of 1,3-diricinoleoyl-2-oleoyl-sn-glycerols (ROR) among the three stereospecific isomers, RRO, ROR and ORR, was about 91%. The contents of other 1,3-diricinoleoyl-2-acyl-glycerols among the three stereospecific isomers were as follows: 1,3-diricinoleoyl-2-linoleoyl-sn-glycerol, 95%; 1,3-diricinoleoyl-2-linolenoyl-sn-glycerol, 96%; 1,3-diricinoleoyl-2-stearoyl-sn-glycerol, 96%; 1,3-diricinoleoyl-2-palmitoyl-sn-glycerol, 78%; and 1,3-diricinoleoyl-2-lesqueroloyl-sn-glycerol, 31%. These non-hydroxyl fatty acids were mostly at the sn-2 position of triacylglycerols in castor oil. These results suggest that phospholipase A2 hydrolysis of phosphatidylcholine (PC) containing non-hydroxyl fatty acid at the sn-2 position is either blocked or partially blocked in vivo. Phospholipase A2 hydrolysis of 2-lesqueroloyl-PC is not blocked and is similar to that of 2-ricinoleoyl-PC. Transgenic inhibition of phospholipase C hydrolysis of PC might be used to block the incorporation of non-hydroxyl fatty acids into triacylglycerols, thus increasing the content of ricinoleate in seed oil.     

Determination of the predominant catechins in Acacia catechu by liquid chromatography/electrospray ionization-mass spectrometry

A high-performance liquid chromatography coupled with electrospray ionization mass spectrometry (LC/ESI-MS) method under selected ion monitoring mode (SIM) was developed to quantitate the predominant catechins, catechin, epicatechin, epicatechin-3-O-gallate, and epigallocatechin-3-O-gallate, in the medicinal plant catechu (Acacia catechu). Other major secondary products including caffeine, flavanol dimers, and flavonol glycosides were also identified by their molecular ion peaks and fragmentation peaks using LC/MS and LC/MS/MS. For the investigated ion concentration ranges of catechin, epicatechin, epicatechin-3-O-gallate, and epigallocatechin-3-O-gallate, good linearities (r2 > 0.99) were obtained for each calibration curve. Validation for this method showed an accuracy ranging from 1.06 to 11.76%, and the precision (relative standard deviation) varied between 1.60 and 9.36% for these four analytes. This is the first quantitative determination of all predominant catechins in catechu heartwood and leaves.     

Arabica and robusta coffees: identification of major polar compounds and quantification of blends by direct-infusion electrospray ionization-mass spectrometry

Considering that illegal admixture of robusta coffee into high-quality arabica coffee is an important task in coffee analysis, we evaluated the use of direct-infusion electrospray ionization-mass spectrometry (ESI-MS) data combined with the partial least-squares (PLS) multivariate calibration technique as a fast way to detect and quantify arabica coffee adulterations by robusta coffee. A total of 16 PLS models were built using ESI± quadrupole time-of-flight (QTOF) and ESI± Fourier transform ion cyclotron resonance (FT-ICR) MS data from hot aqueous extracts of certified coffee samples. The model using the 30 more abundant ions detected by ES+ FT-ICR MS produced the most accurate coffee blend percentage prediction, and thus, it was later successfully employed to predict the blend composition of commercial robusta and arabica coffee. In addition, ESI± FT-ICR MS analysis allowed for the identification of 22 compounds in the arabica coffee and 20 compounds in the robusta coffee, mostly phenolics.     

Analysis of organophosphorus pesticides in honeybee by liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry

Pesticides applied in extended agricultural fields may be controlled by means of bioindicators, such as honeybees, in which are the pesticides bioaccumulate. Liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry (LC-APCI-MS) experiments with positive (PI) and negative (NI) ion modes were optimized for the analysis of 22 organophosphorus pesticides in honeybee samples. The extraction required 3 g of sample, which was extracted with acetone. The extract was purified with coagulating solution and reextracted with Cl(2)CH(2). Pesticides studied could be detected by both ionization modes except for parathion, parathion-methyl, and bromophos, which did not give signals in PI mode, and triazophos, which was not detected in NI mode. Fragmentation voltage and vaporizer temperature were optimized to achieve the highest sensitivity. The spectra profile of each pesticide in PI mode showed the [M + H](+) ion as the main signal, whereas in NI mode only fragment ions were shown. The detection limit obtained in selected ion monitoring mode ranged from 1 to 15 microg kg(-1). The average recoveries from spiked honeybees at various concentration levels (0.5-5 mg kg(-1)) exceeded 65% with relative standard deviations of 4-15%. The method was applied to real samples, in which residues of coumaphos and dimethoate were detected.     

Determination of linear response in the detection of aroma compounds by atmospheric pressure ionization-mass spectrometry (API-MS)

Linearity and detection thresholds of atmospheric pressure ionization-mass spectrometry (API-MS) were determined for 11 aroma compounds in air at concentrations ranging from 50 ppb to approximately 450 ppm (moles of volatile per mole of air). In most cases, the protonated molecular ion (i.e., m/z = M + 1) was the base peak throughout the range; however, some compounds showed an increase in fragmentation at lower concentrations. Detection limits varied greatly (from 50 ppb to 14 ppm) depending upon the aroma compound being measured. The linear range was also strongly dependent upon the aroma compound, with values ranging from <10-fold change in concentration to >4000-fold change in concentration depending upon the volatile being studied. The two volatiles with poor detection thresholds also exhibited the smallest linear range. Most compounds had linear ranges of >200. There was no apparent relationship between gas-phase basicity and either detection limit or linear range.     

Analysis of limonoid glucosides from citrus by electrospray ionization liquid chromatography-mass spectrometry

An electrospray ionization liquid chromatography-mass spectrometry (ESI-LC-MS) method for the detection and quantitation of limonoid glucosides has been developed. Negative ions [M - H(+)](-) characteristic of six limonoid glucosides can be detected and quantified from selected ion monitoring chromatograms using carminic acid as an internal standard. The described method has been applied to the analysis of limonoid glucoside content in various liquid and solid Citrus spp. samples as well as complex mixtures of partially purified limonoid glucosides. Rapid and sensitive qualitative screening of samples for limonoid glucosides can also be accomplished with slight modifications of the method.     

Direct aqueous injection liquid chromatography/electrospray ionization-mass spectrometry/mass spectrometry analysis of water for atrazine, simazine, and their chlorotriazine metabolites

A method is reported for the determination of atrazine, simazine, and their respective dealkylated chlorotriazine metabolites in ground, surface, and finished drinking water. Water samples are diluted 1:4 in an injection vial prior to analysis using liquid chromatography/electrospray ionization-mass spectrometry/mass spectrometry (LC/ESI-MS/MS). The lower limit of method validation is 0.10 microg/L (ppb) for 2-chloro-4-(ethylamino)-6-isopropylamino)-s-triazine (atrazine, G-30027), 2-chloro-4, 6-(diethylamino)-s-triazine (simazine, G-27692), 2-amino-4-chloro-6-(isopropylamino)-s-triazine (deethylatrazine, DEA, or G-30033), 2-amino-4-chloro-6-(ethylamino)-s-triazine (deisopropylatrazine, DIA, or G-28279), and 2,4-diamino-6-chloro-s-triazine (didealkylatrazine, DDA, or G-28273). The overall mean procedural recoveries (and % relative standard deviations) for atrazine, simazine, DEA, DIA, and DDA are 98 (4.4), 102 (3.6), 99 (4.8), 103 (4.0), and 109% (4.8%), respectively, in finished drinking water; 108 (2.7), 104 (5.4), 113 (4.5), 111 (5.2), and 105% (5.3%), respectively, in groundwater; and 96 (6.9), 103 (4.2), 102 (4.4), 102 (5.2), and 102% (8.2%), respectively, in surface water. The method validation was conducted under U.S. EPA FIFRA Good Laboratory Practice Guidelines 40 CFR 160.     

Determination of N-(carboxymethyl)fumonisin B(1) in corn products by liquid chromatography/electrospray ionization--mass spectrometry

It is well-known that fumonisin B(1) (FB(1)) in corn meal decreases during baking, frying, and cooking, but it is still not exactly clear how heating affects the formation of N-(carboxymethyl)fumonisin B(1) (NCM-FB(1)), the reaction product of FB(1) and reducing sugars. In model experiments corn grits were spiked with FB(1) (2 mg/kg) and D-glucose (50 g/kg) or sucrose (50 g/kg) and manufactured into extrusion products at various temperatures (160--180 degrees C) and moisture levels (16--20%). A liquid chromatography/electrospray ionization-mass spectrometry method using isotopically labeled fumonisin FB(1)-d(6) as an internal standard was developed for the determination of NCM-FB(1). For sample cleanup solid-phase C18 cartridges were used. The detection limit achieved with this method was 10 ng/g (signal-noise ratio = 3:1) using the protonated molecule [M + H](+) signal of NCM-FB(1) (m/z 780) in the selected ion monitoring mode. Low concentrations of NCM-FB(1) (29-97 ng/g) were detected in all samples spiked with D-glucose and FB(1), whereas those spiked with FB(1) and sucrose showed only NCM-FB(1) in samples produced at 180 degrees C (NCM-FB(1) = 27 ng/g). Various corn-containing food samples from the German market were analyzed for the presence of NCM-FB(1), FB(1), and hydrolyzed fumonisin B(1) (HFB(1)). All samples were contaminated with FB(1) (22--194 ng/g) and HFB(1) (5--247 ng/g). Six of nine samples contained NCM-FB(1) in low concentrations ranging from 10 to 76 ng/g. From these data and the low toxicity of NCM-FB(1) it can be concluded that the significance of NCM-FB(1) in food seems to be a minor one.     

Monitoring glycation of lysozyme by electrospray ionization mass spectrometry

Electrospray ionization mass spectrometry (ESI-MS) was used to study the glycation of lysozyme by D-glucose (LZM-G) and by D-fructose (LZM-F) under dry heating conditions in the presence and in the absence of oxygen. ESI-MS proved to be a precise method for monitoring protein glycation with respect to following the extent of glycation and changes in the glycoconjugate profile with time. The ESI-MS spectrum of glycated LZM revealed a heterogeneous distribution of glycoforms of LZM at different reaction stages. D-Glucose showed a higher level of reactivity with the amino groups of LZM than D-fructose, both in the presence and in the absence of oxygen. The presence of oxygen in the reaction system induced oxidative side reactions, which competed with and slowed the initial rate of formation of Amadori or Heyns products. The more reactive glycoxidation products formed during the initial stages of incubation in the presence of oxygen accelerated the rate of glycation during the later stages of incubation and increased the involvement of arginine residues of LZM in the glycation reaction. The interaction between the initial glycoxidation product(s) of the reducing sugars and intact lysozyme during the later stages of incubation was observed by the appearance of a different cluster of glycoconjugates in the mass spectrum during the latter stages of incubation. The molecular weight differences between the molecular ions of the new cluster of glycoconjugates are consistent with the formation of D-glucosone from the autoxidation of D-glucose or from the oxidative cleavage of the glucose-lysozyme imine adduct in the lysozyme-glucose system. The effect of oxygen-induced glycoxidation on the glycation reaction was also more pronounced in the LZM-G system compared with that in the LZM-F system.     

Analysis of Flavan-3-ols and procyanidins in food samples by reversed phase high-performance liquid chromatography coupled to electrospray ionization tandem mass spectrometry (RP-HPLC-ESI-MS/MS)

Concentrations of the main dimeric and trimeric procyanidins (PC) and their monomeric constitutive units catechin (CT) and epicatechin (EC) were determined in food samples by using reversed phase high-performance liquid chromatography coupled to electrospray ionization tandem mass spectrometry (RP-HPLC-ESI-MS/MS). In a first step, 12 PCs (PC B1, B2, B3, B4, B5, B6, B7, B8, C1, C2, and A2 and cinnamtannin B1), of which most are not commercially available, were isolated from plant materials or synthesized and purified by a combination of column chromatographic separation techniques with different stationary phases. These PCs in combination with CT and EC were used as standard substances for identification and quantification during the following screening of food samples by RP-HPLC-ESI-MS/MS analysis. The main focus of the newly developed RP-HPLC-ESI-MS/MS method is the compensation of matrix effects by using the echo-peak technique simulating internal standard injection. The suitability of this new method was demonstrated by the determination of recovery rates being 90% or higher. Use of this method allowed the determination of patterns and concentrations of PCs in 55 food samples.     

Simultaneous identification of soyasaponins and isoflavones and quantification of soyasaponin Bb in soy products,using liquid chromatography/electrospray ionization-mass spectrometry

A method has been developed for simultaneous identification of soyasaponins and soy isoflavones in soy products, based on liquid chromatography/electrospray ionization-mass spectrometry (LC/ESI-MS). Soy-based nutraceutical products were analyzed by LC/ESI-MS with detection of protonated and sodiated molecular ions, as well as characteristic fragment ions for these compounds. Soy isoflavones were characterized by a strong protonated molecular ion in addition to corresponding [aglycone + H](+) ions. Monitoring the soyasaponin-specific protonated aglycone and dehydrated aglycone ions throughout the chromatogram provided a unique fingerprint for soyasaponin content in the samples. This mass spectrometric fingerprint also allowed immediate classification of the soyasaponin analytes as group A or B soyasaponins, based on the unique masses of aglycone ions observed for each class. Quantification of soyasaponin B(b) in soy-derived materials, based on the use of a purified soyasaponin B(b) standard and a glycyrrhizin internal standard, has been accomplished.     

Authentication of animal fats using direct analysis in real time (DART) ionization-mass spectrometry and chemometric tools

A combination of direct analysis in real time (DART) ionization coupled to time-of-flight mass spectrometry (TOFMS) and chemometrics was used for animal fat (lard and beef tallow) authentication. This novel instrumentation was employed for rapid profiling of triacylglycerols (TAGs) and polar compounds present in fat samples and their mixtures. Additionally, fat isolated from pork, beef, and pork/beef admixtures was analyzed. Mass spectral records were processed by principal component analysis (PCA) and stepwise linear discriminant analysis (LDA). DART-TOFMS profiles of TAGs were found to be more suitable for the purpose of discrimination among the examined fat types as compared to profiles of polar compounds. The LDA model developed using TAG data enabled not only reliable classification of samples representing neat fats but also detection of admixed lard and tallow at adulteration levels of 5 and 10% (w/w), respectively. The presented approach was also successfully applied to minced meat prepared from pork and beef with comparable fat content. Using the DART-TOFMS TAG profiles of fat isolated from meat mixtures, detection of 10% pork added to beef and vice versa was possible.     

Rapid evaluation of phenolic component profile and analysis of oleuropein aglycon in olive oil by atmospheric pressure chemical ionization-mass spectrometry (APCI-MS)

Epidemiological studies have linked the Mediterranean diet with a low incidence of cardiovascular diseases. Olive oil, the major fat component of this diet, is characterized by antioxidant properties related to their content in catecholic components, particularly oleuropein aglycon. Therefore quantification of these components in edible oils may be important in determining the quality, and consequently its commercial value. The present method allows us to obtain the profile of the phenolic components of the oil from the methanolic extracts of the crude olive oil. In particular tyrosol, hydroxytyrosol, elenolic acid, deacetoxyligstroside and deacetoxyoleuropein aglycons, ligstroside and oleuropein aglycons, and 10-hydroxy-oleuropein are clearly identified by atmospheric pressure chemical ionization-mass spectrometry (APCI-MS). Moreover, oleuropein and its isomers present in the oil are quantified by APCI-MS/MS analysis of the extracts without preliminary separation from other phenolic compounds.     

Analysis of patulin in pear- and apple-based foodstuffs by liquid chromatography electrospray ionization tandem mass spectrometry

A liquid chromatography electrospray ionization tandem mass spectrometry method for the determination of patulin in apple- and pear-based foodstuffs was developed. The sample preparation is based on the QuEChERS procedure involving an initial extraction step with water and acetonitrile, followed by a partitioning step after the addition of magnesium sulfate and sodium chloride. The cleanup was performed by using dispersive solid-phase extraction with a mixture of magnesium sulfate, primary secondary amine sorbent, and n-octadecylsiloxane sorbent added together to the extract. The cleaned extract was finally evaporated and reconstituted in water prior to injection. Quantitation was performed by isotope dilution using ((13)C(7))-patulin as internal standard. The method was first fully validated in three different baby food products including apple-pear juice, apple-pear puree, and infant cereals. Then the scope of application of the method was extended to pear concentrate, raw apples, apple flakes (naturally contaminated), dried apples, and yogurt. The sensitivity achieved by the method in all matrices gave limits of detection (LOD) and quantitation (LOQ) of ≤0.5 and ≤10 μg/kg, respectively, which was compliant with maximum levels settled in Commission Regulation (EC) No. 1881/2006. Method performances for all matrices also fulfilled the criteria established in the CEN/TR 16059:2010 document. Indeed, recoveries were within the 94-104% range; relative standard deviations of repeatability (RSD(r)) and intermediate reproducibility (RSD(IR)) were ≤7.5 and ≤13.0%, respectively, and trueness in an infant apple drink (FAPAS 1642) was measured at 99%.     

Analysis of isomeric forms of oxidized triacylglycerols using ultra-high-performance liquid chromatography and tandem mass spectrometry

Detailed studies on the regioisomeric structures of oxidized species of triacylglycerols (TAG), formed in food during storage and processing, have not been published thus far. In this study, an analytical approach based on efficient ultra-high-performance liquid chromatographic (UHPLC) separation of different isomers of oxidized TAG species and their tandem mass spectrometric analysis was created. A linear solvent gradient based on acetonitrile and acetone was used in the UHPLC method. A novel method utilizing positive ion ESI using ammonia supplemented in the nebulizer gas was used to produce ammonium adduct ions for mass spectrometric analysis. With the UHPLC method used, different regioisomers of TAG species containing oxidized linoleic or oleic acid could be efficiently resolved. Differences in the fragmentation patterns of many of the oxidized TAG isomers could be demonstrated by the tandem mass spectrometric method. On the basis of the results, the approach enables regiospecific analysis of oxidized TAG molecules.     

Pharmacokinetic study of nobiletin and tangeretin in rat serum by high-performance liquid chromatography-electrospray ionization-mass spectrometry

Nobiletin (NOB) and tangeretin (TAN), two of the main polymethoxylated flavones (PMFs) in citrus, influence a number of key biological pathways in mammalian cells. Although the impacts of NOB and TAN on glucose homeostasis and cholesterol regulation have been investigated in human clinical trials, much information is still lacking about the metabolism and oral bioavailability of these compounds in animals. In this study, NOB and TAN were administered to rats by gavage and intraperitoneal (ip) injection, and the blood serum concentrations of these compounds and their main metabolites were monitored by high-performance liquid chromatography-electrospray ionization-mass spectrometry (HPLC-ESI-MS). In addition to the administered compounds, two metabolites of TAN and eight metabolites of NOB were detected and measured over 24 h. With identical oral doses, nearly 10-fold higher absorption of NOB occurred compared to TAN. For both compounds, maximum levels of glucuronidated metabolites occurred in the blood serum at later time points (～5-8 h) compared to the earlier T(max) values for NOB and TAN. In most cases the glucuronides occurred at substantially higher concentrations than the aglycone metabolites. Low levels of NOB and TAN and their metabolites were detectable in rat blood serum even at 24 h after treatment.     

Simultaneous determination of fumonisin B(1) and hydrolyzed fumonisin B(1) in corn products by liquid chromatography/electrospray ionization mass spectrometry

A method for the simultaneous determination of fumonisin B(1) (FB(1)) and its major hydrolysis product (HFB(1)), which is known to be formed during alkaline treatment of fumonisin-containing corn meal, was devised to analyze the levels of these mycotoxins in corn products available on the German market. Liquid chromatography/electrospray mass spectrometry in combination with selected ion monitoring (SIM) was used for unambiguous detection of FB(1) and HFB(1) after extraction of samples with acetonitrile/methanol/water (25:25:50) and solid-phase C18 cleanup. Quantitation was carried out using labeled fumonisin FB(1)-D(6) as an internal standard. The detection limits achieved with this method were 8 ng/g for HFB(1) (signal-noise ratio = 5:1) and 5 ng/g for FB(1) (s/n = 5:1) using the protonated molecule signals m/z 406 and 722 in the SIM mode. A screening of several corn-containing foodstuffs, among them extrusion products and alkali-processed corn food such as tortilla chips, showed HFB(1) and FB(1) contamination with levels of 8-80 and 5-450 ng/g, respectively.     

Determination of carbamate pesticide residues in vegetables and fruits by liquid chromatography-atmospheric pressure photoionization-mass spectrometry and atmospheric pressure chemical ionization-mass spectrometry

A liquid chromatography-atmospheric pressure photoionization (APPI)-mass spectrometry method was developed for the determination of 22 carbamates including their metabolites in vegetables and fruits. For the optimization of APPI, several APPI ion source parameters were examined. As a result, many carbamates with APPI using the optimized parameter gave simple mass spectra, and a strong signal corresponding to [M + H](+) was observed except for aldicarb. However, some carbamate metabolites gave ammonium adduct ions [M + NH(4)](+) as base peak ions. The mean recovery of each carbamate from grape and onion samples spiked at 5 ng/g was 81.7-105.7%, with relative standard deviations of 3.3-5.9%. Furthermore, matrix constituents did not significantly influence the ionization efficiency. The limit of detection (S/N = 3) in grape and onion was in the range of 0.33-3.33 ng/g. For the robustness of this method, this system has been used to analyze 50 samples, and the intensities for all carbamates were found to be unaffected by the contamination of the APPI source by sample matrix constituents. This result indicates that the method is reliable.     

Analysis of bitter limonoids in citrus juices by atmospheric pressure chemical ionization and electrospray ionization liquid chromatography-mass spectrometry

Improved analytical techniques for bitter limonoids in citrus and citrus juices can expedite the evaluation of freeze-induced citrus damage for citrus growers and juice quality for citrus juice producers. Microbore normal-phase and reverse-phase chromatography coupled to a mass spectrometer operating in a positive ion atmospheric pressure chemical ionization and electrospray ionization modes were found to be rapid, selective, and sensitive methods for the analysis of the bitter limonoids limonin and nomilin in citrus juices. Analysis was performed on a chloroform extract of citrus juice to which an internal standard was added. The methods are capable of detecting citrus limonoids in citrus juice in the 60-200 picogram range and quantifying citrus juice limonoids in concentrations as low as 120 picograms. An accurate "total limonoid bitterness" in citrus juice, as represented by the combined occurrence of limonin and nomilin, is easily determined by these methods.     

Characterization of cigarette tobacco by direct electrospray ionization-ion trap mass spectrometry (ESI-ITMS) analysis of the aqueous extract--a novel and simple approach

In support of the efforts to combat smuggling, as well as illegal sale and distribution of cigarettes, an analytical approach for the characterization of tobacco has been proposed and evaluated. It involves aqueous extraction of the filler tobaccos followed by direct analysis of the extracts by electrospray ionization-ion trap mass spectrometry (ESI-ITMS) in the negative mode. Typically, the deprotonated ions, [M - H](-), of organic acids (malic, citric, caffeic, quinic acid) and polyphenols (chlorogenic acid, rutin, scopoletin) were detected. MS/MS spectra of the ion at m/z 191, which is the [M - H](-) of quinic acid, citric acid, and scopoletin, and a fragment ion of chlorogenic acid were acquired. Significant differences in the MS and MS/MS spectra were observed between counterfeit samples and the corresponding authentic brand name cigarettes. Analysis of 25 commercial cigarettes showed that straight Virginia blends were readily distinguished from the blended products containing different tobacco types (Virginia, burley, and Oriental). The former exhibited consistently higher relative abundances of m/z 353 (chlorogenic acid) to m/z 133 (malic acid) in the MS spectra (0.9-1.2 vs 0.4-0.6) and higher intensity ratios of m/z 176 (scopoletin) to m/z 173 (0.4-0.8 vs 0.1-0.3) and of m/z 127 (quinic acid) to m/z 173 (0.7-1.0 vs 0.3-0.5) in the MS/MS spectra. Evidence is presented to demonstrate that the spectral differences were related not only to the tobacco type (Virginia, burley and Oriental) but also to the tobacco part (stem, lamina) used in the manufacture of the cigarettes.