Aloperine alleviates lipopolysaccharide-induced acute lung injury by inhibiting NLRP3 inflammasome activation.

RATIONALE: Excessive activation of the NLRP3 inflammasome is involved in the pathological progression of acute lung injury (ALI). Aloperine (Alo) has anti-inflammatory effects in many inflammatory disease models; however, its role in ALI remains elusive. In this study, we addressed the role of Alo in NLRP3 inflammasome activation in both ALI mice and LPS-treated RAW264.7 cells. METHODS: The activation of the NLRP3 inflammasome in LPS-induced ALI lungs was investigated in C57BL/6 mice. Alo was administered in order to study its effect on NLRP3 inflammasome activation in ALI. RAW264.7 cells were used to evaluate the underlying mechanism of Alo in the activation of the NLRP3 inflammasome in vitro. RESULTS: The activation of the NLRP3 inflammasome occurs in the lungs and RAW264.7 cells under LPS stress. Alo attenuated the pathological injury of lung tissue as well as downregulates the mRNA expression of NLRP3 and pro-caspase-1 in ALI mice and LPS-stressed RAW264.7 cells. The expression of NLRP3, pro-caspase-1, and caspase-1 p10 were also significantly suppressed by Alo in vivo and in vitro. Furthermore, Alo decreased IL-1ß and IL-18 release in ALI mice and LPS-induced RAW264.7 cells. In addition, ML385, a Nrf2 inhibitor, weakened the activity of Alo, which inhibited the activation of the NLRP3 inflammasome in vitro. CONCLUSION: Alo reduces NLRP3 inflammasome activation via the Nrf2 pathway in ALI mice.

Application of Biotechnology in Specific Spoilage Organisms of Aquatic Products.

Aquatic products are delicious and have high nutritive value, however, they are highly perishable during storage due to the growth and metabolism of microorganisms. The spoilage process of aquatic products was demonstrated to be highly related to the composition of microorganisms, in which the specific spoilage organisms (SSOs) are the main factors. In this article, the spoilage indicators of SSOs were systematically described, which could make a comprehensive evaluation of the quality of aquatic products. Quorum sensing (QS) regulates the growth, metabolism and characteristics of SSOs, the common signaling molecules and the QS system in the major SSOs of aquatic products were discussed. Moreover, we compared various technologies for the analysis of SSOs in aquatic products. Besides, quality control techniques based on microbiota regulating of aquatic products, including physical, chemical and biological preservation strategies, were also compared. In conclusion, novel preservation technologies and hurdle techniques are expected to achieve comprehensive inhibition of SSOs.

CRISPR/Cas tools for enhancing the biopreservation ability of lactic acid bacteria in aquatic products.

Lactic acid bacteria (LAB) plays a crucial role in aquatic products biopreservation as it can inhibit many bacteria, in particular the specific spoilage organisms (SSOs) of aquatic products, by competing for nutrients or producing one or more metabolites which have antimicrobial activity, such as bacteriocins. Lactobacillus spp. and Lactococcus spp. are the most commonly used Lactic acid bacterias in aquatic products preservation. The improvement of gene editing tools is particularly important for developing new lactic acid bacteria strains with superior properties for aquatic products biopreservation. This review summarized the research progress of the most widely used CRISPR/Cas-based genome editing tools in Lactobacillus spp. and Lactococcus spp. The genome editing tools based on homologous recombination and base editor were described. Then, the research status of CRISPRi in transcriptional regulation was reviewed briefly. This review may provide a reference for the application of CRISPR/Cas-based genome editing tools to other lactic acid bacteria species.

Supercritical Fluid CO2 Extraction and Microcapsule Preparation of Lycium barbarum Residue Oil Rich in Zeaxanthin Dipalmitate.

The scope of this investigation aimed at obtaining and stabilizing bioactive products derived from Lycium barbarum seeds and peels, which were the byproducts in the processing of fruit juice. Zeaxanthin dipalmitate is a major carotenoid, comprising approximately 80% of the total carotenoid content in the seeds and peels. The method of obtainment was supercritical fluid CO2 extraction, studying different parameters that affect the oil yield and content of zeaxanthin dipalmitate. The optimized protocol to enact successful supercritical fluid CO2 extraction included optimum extraction pressure of 250 bar, temperature at 60 °C over a time span of 2.0 h, and a CO2 flow of 30 g/min, together with the use of a cosolvent (2% ethanol). The yields of oil and zeaxanthin dipalmitate under these optimal conditions were 17 g/100 g and 0.08 g/100 g, respectively. The unsaturated fatty acids were primarily linoleic acid (C18:2), oleic acid (C18:1), and γ-linolenic acid (C18:3), with their contents being as high as 91.85 ± 0.27% of the total fatty acids. The extract was a red-colored oil that was consequently microencapsulated through spray-drying with octenylsuccinate starch, gum arabic, and maltodextrin (13.5:7.5:3, w/w) as wall materials to circumvent lipid disintegration during storage and add to fruit juice in a dissolved form. The mass ratio of core material and wall material was 4:1. These materials exhibited the highest microencapsulation efficiency (92.83 ± 0.13%), with a moisture content of 1.98 ± 0.05% and solubility of 66.22 ± 0.24%. The peroxide content level within the microencapsulated zeaxanthin dipalmitate-rich oil remained at one part per eight in comparison to the unencapsulated oil, following fast-tracked oxidation at 60 °C for 6 weeks. This indicated the potential oxidation stability properties of microcapsule powders. Consequently, this microencapsulated powder has good prospects for development, and can be utilized for a vast spectrum of consumer health and beauty products.

Character changes and Transcriptomic analysis of a cassava sexual Tetraploid.

BACKGROUND: Cassava (Manihot esculenta Crantz) is an important food crop known for its high starch content. Polyploid breeding is effective in its genetic improvement, and use of 2n gametes in sexual polyploid breeding is one of the potential methods for cassava breeding and improvement. In our study, the cassava sexual tetraploid (ST), which carries numerous valuable traits, was successfully generated by hybridizing 2n female gametes SC5 (♀) and 2n male gametes SC10 (♂). However, the molecular mechanisms remain unclear. To understand these underlying molecular mechanisms behind the phenotypic alterations and heterosis in ST plants, we investigated the differences in gene expression between polyploids and diploids by determining the transcriptomes of the ST plant and its parents during the tuber root enlargement period. We also compared the characters and transcriptomes of the ST plant with its parents. RESULTS: The ST plant was superior in plant height, stem diameter, leaf area, petiole length, plant weight, and root weight than the parent plants, except the leaf number, which was lower. The number of starch granules was higher in the roots of ST plants than those in the parent plants after five months (tuber root enlargement period), which could be due to a higher leaf net photosynthetic rate leading to early filling of starch granules. Based on transcriptome analysis, we identified 2934 and 3171 differentially expressed genes (DEGs) in the ST plant as compared to its female and male parents, respectively. Pathway enrichment analyses revealed that flavonoid biosynthesis and glycolysis/gluconeogenesis were significantly enriched in the ST plants, which might contribute to the colors of petiole (purple-red), root epidermis (dark brown), and tuber starch accumulation, respectively. CONCLUSIONS: After sexual polyploidization, the phenotype of ST has changed significantly in comparison to their diploid parents, mainly manifest as enlarged biomass, yield, early starch filling, deep colored petiole and root epidermis. The tetraploid plants were also mature early due to early starch grain filling. Owing to enriched flavonoid biosynthesis and glycolysis/gluconeogenesis, they are possibly resistant to adversity stresses and provide better yield, respectively.

MeABL5, an ABA Insensitive 5-Like Basic Leucine Zipper Transcription Factor, Positively Regulates MeCWINV3 in Cassava (Manihot esculenta Crantz).

The basic leucine zipper (bZIP) transcription factor family plays crucial roles in multiple biological processes, especially stress responses. Cassava (Manihot esculenta Crantz) is an important tropical crop with a strong tolerance to environmental stresses such as drought, heat, and low-fertility environments. Currently, limited information is available regarding the functional identification of bZIP transcription factors in response to abiotic stress in cassava. Herein, a gene encoding an ABA Insensitive 5 (ABI5)-like transcription factor, designated as MeABL5, was identified in cassava. Sequence and phylogenetic analysis showed that MeABL5 is a cassava bZIP transcription factor that is not included in the previously identified cassava bZIP family members, belongs to subfamily A, and has high sequence similarity to ABI5-like proteins. Subcellular localization and transactivation assays revealed that MeABL5 was a nuclear-localized protein and possessed transactivation activity. Furthermore, MeABL5 was able to specifically interact with the ABRE cis-element in the promoter of the cassava major cell wall invertase gene, MeCWINV3, in vitro and in vivo. MeABL5 and MeCWINV3 exhibited similar expression patterns in various organs or tissues and under abiotic stress in cassava. The expressions of MeABL5 and MeCWINV3 within cassava plantlets were both induced by exogenous abscisic acid (ABA), gibberellic acid (GA3), methyl jasmonate (MeJA), and heat. Overexpression of MeABL5 increased the activity of the MeCWINV3 gene, and the up-regulated expressions of MeCWINV3 were significantly activated under ABA-, salicylic acid (SA)-, and MeJA-induced conditions. Overall, these results suggest that MeABL5 is a positive regulator of MeCWINV3 and might participate in the robust resistance of cassava in response to abiotic stress. This study also provides a foundation for further research on ABA-mediated and stress-related signaling pathways in cassava.

Isolation and Characterization of Ftsz Genes in Cassava.

The filamenting temperature-sensitive Z proteins (FtsZs) play an important role in plastid division. In this study, three FtsZ genes were isolated from the cassava genome, and named MeFtsZ1, MeFtsZ2-1, and MeFtsZ2-2, respectively. Based on phylogeny, the MeFtsZs were classified into two groups (FtsZ1 and FtsZ2). MeFtsZ1 with a putative signal peptide at N-terminal, has six exons, and is classed to FtsZ1 clade. MeFtsZ2-1 and MeFtsZ2-2 without a putative signal peptide, have seven exons, and are classed to FtsZ2 clade. Subcellular localization found that all the three MeFtsZs could locate in chloroplasts and form a ring in chloroplastids. Structure analysis found that all MeFtsZ proteins contain a conserved guanosine triphosphatase (GTPase) domain in favor of generate contractile force for cassava plastid division. The expression profiles of MeFtsZ genes by quantitative reverse transcription-PCR (qRT-PCR) analysis in photosynthetic and non-photosynthetic tissues found that all of the MeFtsZ genes had higher expression levels in photosynthetic tissues, especially in younger leaves, and lower expression levels in the non-photosynthetic tissues. During cassava storage root development, the expressions of MeFtsZ2-1 and MeFtsZ2-2 were comparatively higher than MeFtsZ1. The transformed Arabidopsis of MeFtsZ2-1 and MeFtsZ2-2 contained abnormally shape, fewer number, and larger volume chloroplasts. Phytohormones were involved in regulating the expressions of MeFtsZ genes. Therefore, we deduced that all of the MeFtsZs play an important role in chloroplast division, and that MeFtsZ2 (2-1, 2-2) might be involved in amyloplast division and regulated by phytohormones during cassava storage root development.

Identification, Expression, and Functional Analysis of the Fructokinase Gene Family in Cassava.

Fructokinase (FRK) proteins play important roles in catalyzing fructose phosphorylation and participate in the carbohydrate metabolism of storage organs in plants. To investigate the roles of FRKs in cassava tuber root development, seven FRK genes (MeFRK1-7) were identified, and MeFRK1-6 were isolated. Phylogenetic analysis revealed that the MeFRK family genes can be divided into α (MeFRK1, 2, 6, 7) and ß (MeFRK3, 4, 5) groups. All the MeFRK proteins have typical conserved regions and substrate binding residues similar to those of the FRKs. The overall predicted three-dimensional structures of MeFRK1-6 were similar, folding into a catalytic domain and a ß-sheet ''lid" region, forming a substrate binding cleft, which contains many residues involved in the binding to fructose. The gene and the predicted three-dimensional structures of MeFRK3 and MeFRK4 were the most similar. MeFRK1-6 displayed different expression patterns across different tissues, including leaves, stems, tuber roots, flowers, and fruits. In tuber roots, the expressions of MeFRK3 and MeFRK4 were much higher compared to those of the other genes. Notably, the expression of MeFRK3 and MeFRK4 as well as the enzymatic activity of FRK were higher at the initial and early expanding tuber stages and were lower at the later expanding and mature tuber stages. The FRK activity of MeFRK3 and MeFRK4 was identified by the functional complementation of triple mutant yeast cells that were unable to phosphorylate either glucose or fructose. The gene expression and enzymatic activity of MeFRK3 and MeFRK4 suggest that they might be the main enzymes in fructose phosphorylation for regulating the formation of tuber roots and starch accumulation at the tuber root initial and expanding stages.

Structure, Expression, and Functional Analysis of the Hexokinase Gene Family in Cassava.

Hexokinase (HXK) proteins play important roles in catalyzing hexose phosphorylation and sugar sensing and signaling. To investigate the roles of HXKs in cassava tuber root development, seven HXK genes (MeHXK1-7) were isolated and analyzed. A phylogenetic analysis revealed that the MeHXK family can be divided into five subfamilies of plant HXKs. MeHXKs were clearly divided into type A (MeHXK1) and type B (MeHXK2-7) based on their N-terminal sequences. MeHXK1-5 all had typical conserved regions and similar protein structures to the HXKs of other plants; while MeHXK6-7 lacked some of the conserved regions. An expression analysis of the MeHXK genes in cassava organs or tissues demonstrated that MeHXK2 is the dominant HXK in all the examined tissues (leaves, stems, fruits, tuber phloems, and tuber xylems). Notably, the expression of MeHXK2 and the enzymatic activity of HXK were higher at the initial and expanding tuber stages, and lower at the mature tuber stage. Furthermore, the HXK activity of MeHXK2 was identified by functional complementation of the HXK-deficient yeast strain YSH7.4-3C (hxk1, hxk2, glk1). The gene expression and enzymatic activity of MeHXK2 suggest that it might be the main enzyme for hexose phosphorylation during cassava tuber root development, which is involved in sucrose metabolism to regulate the accumulation of starch.

Experiential Avoidance Mediates the Association between Emotion Regulation Abilities and Loneliness.

Experiential avoidance (EA) involves the unwillingness to remain in contact with aversive experiences such as painful feelings, thoughts, and emotions. EA is often associated with the development and maintenance of emotional problems. Since loneliness is characterized by negative emotions such as sadness and pessimism, which is often linked to emotional problems, this study aims to test the mediating effects of EA on the relationship between emotion regulation abilities (ERA) and loneliness in a sample of Chinese adults. Five hundred undergraduates completed questionnaires measuring EA (Acceptance and Action Questionnaire; AAQ-Ⅱ), ERA (Failure-relate action orientation; Action Control Scale, ACS-90) and loneliness (UCLA Loneliness Scale). Structural equation modeling showed that EA fully-mediated the relationship between ERA and loneliness. The findings suggest EA is a key mechanism in explaining why people with high ERA are prone to feeling lower levels of loneliness. In particular, these findings have important implications for designing effective psychological interventions for loneliness.

SpBADH of the halophyte Sesuvium portulacastrum strongly confers drought tolerance through ROS scavenging in transgenic Arabidopsis.

Glycine betaine (GB) accumulation is involved in abiotic stress. However, it is not known whether BADH, the key enzyme of GB synthesis, utilizes the antioxidant system to confer drought stress tolerance. In this study, a novel member of the ALDH10 gene family, SpBADH, was isolated from Sesuvium portulacastrum. The expression of this gene was up-regulated by NaCl, PEG6000, H2O2, ABA and high temperature in S. portulacastrum. SpBADH overexpression in Arabidopsis resulted in higher BADH activity and GB content and might increase tolerance to drought/osmotic stresses, specifically strong tolerance to drought stress. Transgenic lines exhibited lower MDA and H2O2 contents but higher proline, POD, SOD and CAT contents than the wild type under drought and osmotic stresses. SpBADH overexpression in Arabidopsis also enhanced the expression of ROS-related genes including AtSOD, AtPOD, AtCAT, AtAPX and Atpsb under drought and osmotic stresses. Thus, SpBADH increases plant tolerance to drought or osmotic stresses by reducing H2O2, increasing proline, and activating antioxidative enzymes to improve ROS scavenging.

Recombinant expressions of sweet plant protein mabinlin II in Escherichia coli and food-grade Lactococcus lactis.

Sweet plant proteins, which are safe, natural, low-calorie sweeteners, may be suitable replacements for sugars in the food and beverage industries. Mabinlin II, a sweet plant protein, shows the most pronounced heat stability and acid resistance of any of the six known types of plant sweet proteins. However, mabinlin II is difficult to extract from the Capparis masaikai plant, which is itself becoming increasingly scarce. This limits the use of naturally acquired mabinlin II. In this study, recombinant mabinlin II proteins were expressed and purified in Escherichia coli and in food-grade Lactococcus lactis. Recombinant mabinlin II proteins MBL-BH (containing the B-chains of mabinlin II downstream fused with His-tag) and MBL-ABH (containing the A- and B-chains of mabinlin II downstream fused with His-tag) were expressed in E. coli in the form of inclusion bodies. They were then purified and renatured. The refolded MBL-BH was found to be 100 times sweeter than sucrose by weight, but it was not heat-stable. Refolded MBL-ABH was neither sweet nor heat-stable. Recombinant mabinlin II proteins were secreted and expressed intracellularly in food-grade L. lactis, in which the concentrated cell samples and culture medium samples were detected using enzyme-linked immunosorbent assay and Western blotting analysis with anti-mabinlin II polyclonal antibody. This study demonstrated that the single B chain of mabinlin II has a sweet taste. The recombinant mabinlin II proteins have been successfully expressed in food-grade L. lactis, which is a crucial step in the production of mabinlin II through microorganism expression systems.

[Study on chemical changes of salvianolic acid B and lithospermic acid aqueous under conditions of high temperature and high pressure].

OBJECTIVE: To study the chemical changes of salvianolic acid B and lithospermic acid of Salvia miltiorrhiza under the conditions of high temperature and high pressure and explore the reaction mechanism. METHOD: S. miltiorrhiza extracts, salvianolic acid B and lithospermic acid were put in the reactor under the conditions of high temperature and high pressure (120 degrees C, 0.2 MPa), and the chemical changes and stability was studied. RESULT: Salvianolic acid A was the primary product in salvianolic acid B and lithospermic acid's conversion process, and lithospermic acid was an intermediate in the conversion process of salvianolic acid B. Compared with salvianolic acid B, lithospermic acid could convert into more salvianolic acid A and fewer other products in the same conditions. Salvianolic acid A was not stable under the conditions of high temperature and high pressure, and could sequentially convert into other small molecules. CONCLUSION: Referring to the chemical conversion of salvianolic acid B and lithospermic acid, a method of large-scale preparation of salvianolic acid A can be developed.

HPLC/CE-ESI-TOF-MS methods for the characterization of polyphenols in almond-skin extracts.

We have developed two rapid methods using CE and HPLC coupled to ESI-TOF-MS and both these methods have been compared for the separation and characterization of antioxidant phenolic compounds in almond-skin extract. Under optimum CE-ESI-TOF-MS conditions we achieved the determination of nine compounds from the polar fraction in 35 min. Furthermore, by using the HPLC-ESI-TOF-MS method, a total of 23 compounds corresponding to phenolic acids and the flavonoid family were identified from almond skin in only 9 min. The sensitivity, together with mass accuracy and true isotopic pattern of TOF-MS, allowed the identification of a broad series of known phenolic compounds present in almond-skin extracts using HPLC and CE as separation techniques.

Effects of heating on virgin olive oils and their blends: focus on modifications of phenolic fraction.

The phenolic profiles of two different virgin olive oils and their admixtures in different percentages have been analyzed after heating treatments by microwave or conventional oven. Changes in the phenolic profile upon heating were evaluated by chromatographic and spectroscopic methods, also monitoring the antioxidant activity by ABTS(*+) test. 3,4-DHPEA-EA, p-HPEA-EA, and EA showed the highest decreases after thermal treatments. The only compounds that showed a clear increase with heating, in particular by conventional oven, were the dialdehydic form of elenolic acid (EDA) and p-hydroxyphenylethanol linked to the dialdehydic form of elenolic acid (p-HPEA-EDA). A comparison between the variations after heating of the sum of monoaldehydic and dialdehydic forms of phenolic compounds obtained by using different analytical approaches (HPLC-DAD/MSD and 1D and 2D NMR spectroscopy) was made. The results showed a good agreement of these two high-resolution techniques.

Characterization of steroidal saponins in crude extracts from Dioscorea zingiberensis C. H. Wright by ultra-performance liquid chromatography/electrospray ionization quadrupole time-of-flight tandem mass spectrometry.

Steroidal saponins are the major bioactive constituents of Dioscorea zingiberensis C. H. Wright (D. zingiberensis). In this work, ultra-performance liquid chromatography/electrospray ionization quadrupole time-of-flight tandem mass spectrometry (UPLC/Q-TOF-MS/MS) was applied to the separation and characterization of steroidal saponins in crude extracts from D. zingiberensis. The results showed that fragment ions from glycosidic and cross-ring cleavages gave a wealth of structural information related to aglycone skeletons, sugar types and the sequence of sugar units. According to the summarized fragmentation patterns, identification of steroidal saponins from D. zingiberensis could be fulfilled, even when reference standards were unavailable. As a result, a total of thirty-one saponins with five aglycone skeletons, including fourteen new trace saponins, were identified or tentatively elucidated in crude extracts from D. zingiberensis based on their retention times, the mass spectrometric fragmentation patterns, and MS and MS/MS data.

Qualitative screening of phenolic compounds in olive leaf extracts by hyphenated liquid chromatography and preliminary evaluation of cytotoxic activity against human breast cancer cells.

In this work, high-performance liquid chromatography (HPLC) coupled to electrospray time-of-flight mass spectrometry (ESI-TOF-MS) and electrospray ion trap multiple-stage tandem mass spectrometry (ESI-IT-MS(2)) has been applied to screen phenolic compounds in olive leaf extracts. The use of a small particle size C18 column (1.8 micro) provided great resolution and made separation of a lot of isomers possible. The structural characterization was based on accurate mass data obtained by ESI-TOF-MS, and the nature of fragmentation ions were further confirmed by ESI-IT-MS(2) when possible. In addition, we employed tetrazolium salt (MTT)-based assays to assess the effects of olive leaf extracts on the growth of human tumor-derived cells. Upon this approach, we achieved an accurate profile of olive leaf phenolics along with the identification of several important isomers of secoiridoids and flavonoids. This will allow a better understanding of the complete composition of olive-leaf-bioactive compounds as well as their involvement in Olea europaea L. biochemical pathways. Importantly, olive leaf extracts exhibited dose-dependent inhibitory effects on the metabolic status (cell viability) of three breast cancer models in vitro. Since the tumoricidal activity of the extracts should be mainly attributed to the identified olive leaf phenolics, these findings warrant further investigation at the structure-function molecular level to definitely establish the anticancer value of these phytochemicals.

Isopentenyl transferase gene (ipt) downstream transcriptionally fused with gene expression improves the growth of transgenic plants.

This research reports a promising approach to increase a plant's physiological cytokinin content. This approach also enables the increase to play a role in plant growth and development by introducing the ipt gene to downstream transcriptionally fuse with other genes under the control of a CaMV35S promoter, in which the ipt gene is far from the 35S promoter. According to Kozak's ribosome screening model, expression of the ipt gene is reduced by the terminal codon of the first gene and the internal untranslated nucleotides between the fused genes. In the transgenic plants pVKH35S-GUS-ipt, pVKH35S-AOC-ipt, and pVKH35S-AtGolS2-ipt, cytokinins were increased only two to threefold, and the plants grew more vigorously than the pVKH35S-AOC or pVKH35S-AtGolS2 transgenic plants lacking the ipt gene. The vigorous growth was reflected in rapid plant growth, a longer flowering period, a greater number of flowers, more seed product, and increased chlorophyll synthesis. The AOC and AtGolS2 genes play a role in a plant's tolerance of salt or cold, respectively. When the ipt gene transcriptionally fuses with AOC or AtGolS2 in the frame of AOC-ipt and AtGolS2-ipt, slight cytokinin increases were obtained in their transgenic plants; furthermore, those increases played a positive role in improvements of plant growth. Notably, an increased cytokinin volume at the physiological level, in concert with AtGolS2 expression, enhances a plant's tolerance to cold.

CaCl2 enhanced somatic embryogenesis in Manihot esculenta Crantz.

Primary cassava somatic embryos were induced on a medium without CaCl(2), however, no or only a few secondary somatic embryos were formed from them. With 15 mM CaCl(2) in the medium for induction of cassava primary embryos, more secondary somatic embryos were produced from them, and they were much effective in maintaining their embryogenic capacity than the controls of embryos which were induced without CaCl(2).

Tentative characterization of novel phenolic compounds in extra virgin olive oils by rapid-resolution liquid chromatography coupled with mass spectrometry.

Rapid-resolution liquid chromatography (RRLC) coupled with electrospray time-of-flight mass spectrometry (ESI-TOF-MS) and electrospray ion-trap multiple mass spectrometry (ESI-IT-MS(n)) has been applied to separate and characterize the phenolic compounds in five Spanish extra virgin olive oils. The phenolic compounds were characterized on the basis of accurate mass data according to studies by TOF-MS, and the fragmentation ions were further confirmed by IT-MS(n) whenever possible. Eight compounds, hydroxytyrosol (m/z 153), tyrosol (m/z 137), deacetoxy oleuropein aglycon (m/z 319), luteolin (m/z 285), pinoresinol (m/z 357), acetoxypinoresinol (m/z 415), apigenin (m/z 269), and deacetoxy ligstroside aglycon (m/z 303), all normally found in olive oil, were characterized in all five samples. Nevertheless, 30 uncommon phenolic compounds, including 10 ligstroside aglycon isomers (m/z 361) and two methyl oleuropein aglycon isomers (m/z 391), were also characterized in some of the five samples, and their fragmentation pathways were successfully elucidated.