Exon Junction Complex Mediates the Cap-Independent Translation of Circular RNA.

Evidence that circular RNAs (circRNA) serve as protein template is accumulating. However, how the cap-independent translation is controlled remains largely uncharacterized. Here, we show that the presence of intron and thus splicing promote cap-independent translation. By acquiring the exon junction complex (EJC) after splicing, the interaction between circRNA and ribosomes was promoted, thereby facilitating translation. Prevention of splicing by treatment with spliceosome inhibitor or mutating splicing signal hindered cap-independent translation of circRNA. Moreover, EJC-tethering using Cas13 technology reconstituted EJC-dependent circRNA translation. Finally, the level of a coding circRNA from succinate dehydrogenase assembly factor 2 (circSDHAF2) was found to be elevated in the tumorous tissues from patients with colorectal cancer, and shown to be critical in tumorigenesis of colorectal cancer in both cell and murine models. These findings reveal that EJC-dependent control of circSDHAF2 translation is involved in the regulation of oncogenic pathways. IMPLICATIONS: EJC-mediated cap-independent translation of circRNA is implicated in the tumorigenesis of colorectal cancer.

Determination of adulteration, geographical origins, and species of food by mass spectrometry.

Food adulteration, mislabeling, and fraud, are rising global issues. Therefore, a number of precise and reliable analytical instruments and approaches have been proposed to ensure the authenticity and accurate labeling of food and food products by confirming that the constituents of foodstuffs are of the kind and quality claimed by the seller and manufacturer. Traditional techniques (e.g., genomics-based methods) are still in use; however, emerging approaches like mass spectrometry (MS)-based technologies are being actively developed to supplement or supersede current methods for authentication of a variety of food commodities and products. This review provides a critical assessment of recent advances in food authentication, including MS-based metabolomics, proteomics and other approaches.

The identification of soy sauce adulterated with bean species and the origin using headspace solid-phase microextraction coupled with gas chromatography-mass spectrometry.

Soy sauce is one of the significant seasonings in Asia but is often mislabeled in ingredients or substituted with geographical information. With no adequate methods to distinguish the bean sources and the origins of soy sauce, our study designed a seamless headspace solid-phase microextraction coupled with gas chromatography-mass spectrometry (HS-SPME/GC-MS) for analyzing unique volatile components of different soy sauces. Over 400 volatile flavor compounds were identified and the assistance of chemometric analysis successfully discriminated different bean sources (black bean and soybean) and producing regions (Taiwan and Japan). The chemometric models can also perfectly evaluate real samples together with adulterated samples. In brief, these soy sauce volatile signatures can solve the problem of authentication and assist the whole industry in preventing adulteration and producing countries' counterfeit.

Proteomics for species authentication of cod and corresponding fishery products.

Seafood substitutions is a global problem and come under the spotlight in recent years. In Taiwan, Greenland halibut is usually substituted for the cod because of its lower price. Nowadays, DNA technology is widely used for fish species identifications; however, it still has concern about the DNA of processed fishery products might be destroyed. This study was designed to develop a proteomic-based method for fish and fishery product authentication by using ultra performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS/MS) with Sequential window acquisition of all theoretical fragment ion spectra (SWATH). The protein biomarkers from the meat of Alaska pollock, Atlantic cod, and Greenland halibut were identified and validated for species authentication of cod and corresponding fishery products, which might prevent consumer substitutions and fish product mislabeling. Besides, the E. coli proteins can be measured from existing SWATH-MS data though retrospective analysis successfully, it might present the quality of fish meat.

Inhibition of the clinical isolates of Acinetobacter baumannii by Pseudomonas aeruginosa: In vitro assessment of a case-based study.

BACKGROUND: The global rise in nosocomial infections associated with gram-negative bacteria and the spread of multi-drug resistant Acinetobacter baumannii (MDR-AB) pose public health concerns. This study investigates the inhibitory effects and possible inhibitory mechanism of Pseudomonas aeruginosa (PA) on selected clinical strains of A. baumannii (AB) isolated from Taiwanese patients. METHODS: Four and eight clinical strains of AB and PA, respectively, were randomly selected from the bacterial collection of Feng-Yuan Hospital, Taiwan. Antimicrobial-susceptibility was performed on the AB strains. Inhibition potential of the PA strains against AB was assessed by measuring the inhibition zones. In vitro analysis using phenazine-1-carboxamide (PCN) was conducted to assess the possible inhibitory mechanism of PA, which was later confirmed in the clinical isolates by liquid chromatography-mass spectrometry. RESULTS: All the clinical AB strains showed resistance to the eleven antibiotics and were classified as MDR-AB. The nine PA strains exert either a high (PA3596, PA3681, PA3772, and ATCC27853) or a low (PA3613, PA3625, PA3712, PA3715, and PA3744) degree of inhibition against AB strains. 0.25 mg/ml PCN had a clearer inhibition zone than 0.05 mg/ml PCN, suggesting a dose-dependent inhibition of PCN on the AB strains. The four PA strains that demonstrated a high degree of inhibition had a relatively high amount of PCN. CONCLUSION: Selected strains of PA exert inhibitory actions on MDR-AB with PCN being a possible inhibitory agent. This finding raises the possibility of developing effective therapeutic antibiotics and disinfectant from specific components of PA for the treatment and control of Acinetobacter-associated infections in hospital settings.

Metabolic disturbances in systemic lupus erythematosus evaluated with UPLC-MS/MS.

OBJECTIVES: Systemic lupus erythematosus (SLE) is an autoimmune disease. However, no surrogate biomarker is available for SLE diagnosis or predicting disease outcomes. Here, an ultra-performance liquid chromatography coupled with tandem mass spectrometry (UPLC-MS/MS)-based metabolomics strategy was executed to conduct biomarker discovery in SLE. METHODS: Metabolite profiles were analysed using UPLC-MS/MS analysis of serum samples obtained from the discovery cohort. Differentially expressed metabolites were identified using multivariate analyses. During the validation stage, the significant metabolites identified in the discovery cohort were quantified in a validation cohort using multiple reaction monitoring mass spectrometry (MRM-MS). Differences in serum metabolite levels and SLE disease activity markers were examined by using Spearman's correlation analysis. RESULTS: A total of 29 significant metabolites were identified by the UPLC-MS/MS analysis. These metabolites were primarily involved in fatty acid metabolism (20.69%) and phospholipid catabolism (17.24%). In the validation cohort, 11 of 29 metabolites were quantified, which demonstrated increased levels of pyroglutamic acid and L-phenylalanine in SLE patients compared with healthy controls. Patients with lupus nephritis (LN) presented with higher taurine levels, which could serve as a biomarker. The literature review indicated decreased levels of amino acids and adenosine among SLE patients and increased lipids, low-density lipoprotein, and very low-density lipoprotein among LN patients compared to healthy controls. CONCLUSIONS: Fatty acid metabolism and phospholipid catabolism were affected in SLE patients. Pyroglutamic acid and L-phenylalanine have the potential to act as SLE biomarkers, and taurine might be used to distinguish patients with and without LN.

Honey proteomic signatures for the identification of honey adulterated with syrup, producing country, and nectar source using SWATH-MS approach.

Honey is widely consumed by humans, due to its multiple applications as a food constituent and its therapeutic effects. This study reports on the discrimination of honey products from different geographical and botanical sources, as well as honey products containing distinct forms of syrup used in honey adulteration. Sequential window acquisition of all theoretical fragment ion spectra mass spectrometry (SWATH-MS)-based proteomic analysis combined with chemometrics was successfully applied in identifying characteristic proteins that can be used as biomarkers of the original source of honey. Honey samples from different producing regions (Tainan, Changhua, and Taichung), countries (Taiwan and Thailand), and distinct botanical sources (longan and litchi) were clearly distinguished by the developed orthogonal projections to latent structures discriminant analysis (OPLS-DA) model with good fitness and prediction ability. Furthermore, we successfully discriminated the adulteration of honey with syrup in different proportions (even with honey content as low as 20%) with this proteomic SWATH-MS platform.

Variations in bovine milk oligosaccharides after calving using p-aminobenzoic ethyl ester closed-ring labeling and negative ion electrospray LC/MS/MS.

A strategy was proposed to analyze bovine milk oligosaccharides using p-aminobenzoic ethyl ester (ABEE) closed-ring labeling and C18 capillary liquid chromatography negative ion electrospray tandem mass spectrometry. Linkage specific fragment ions were used to identify oligosaccharide isomers. By constructing the mass chromatograms using linkage specific fragment ions, isomers were differentiated based on m/z values as well as temporal separation provided by liquid chromatography. In addition to disialyllactose and the single isomer lacto-N-neohexaose, four pairs of linkage isomers including 3'/6'-sialyllactose (3'/6'-SL), 3'/6'-sialyllactosamine (3'/6'-SLN), 3'/6'-sialylgalactosyl-lactose (3'/6'-SGL), and lacto-N-tetraose/lacto-N-neotetraose (LNT/LNnT) in bovine milk were investigated. Variations of bovine milk oligosaccharides in a lactation period of 72 h after calving were studied. Sialylated oligosaccharide was found to be distinctively more abundant in milk of the first 24 h, decreasing in successive milkings. For the first time, the variation of lacto-N-tetraose in bovine milk was reported.

SWATH-MS-based quantitative proteomics reveals a uniquely intricate defense response in Cnaphalocrocis medinalis-resistant rice.

Cnaphalocrocis medinalis is a major insect pest of rice in Asia. A few defensive enzymes were reported to show higher activities in a resistant rice line (Qingliu) than in a susceptible rice line (TN1) upon leaffolder infestation. However, the overall molecular regulation of the rice defense response against leaffolder herbivory is unknown. Here, differential proteomic analysis by SWATH-MS was performed to identify differentially expressed proteins between the two rice varieties, Qingliu and TN1, at four time points of leaffolder herbivory, 0, 6, 24, and 72 h. Gene Ontology (GO) enrichment of the differentially expressed proteins indicated overrepresentation of (1) photosynthesis, (2) amino acid and derivative metabolic process, and (3) secondary metabolic process. Phenylalanine ammonia lyase and chalcone synthase, which catalyze flavonoid biosynthesis, and lipoxygenase, which catalyzes jasmonic acid biosynthesis, exhibited higher expression in Qingliu than in TN1 even before insect herbivory. Momentary activation of the light reaction and Calvin cycle was detected in Qingliu at 6 h and 24 h of insect herbivory, respectively. At 72 h of insect herbivory, amino acid biosynthesis and glutathione-mediated antioxidation were activated in Qingliu. A defense response involving jasmonic acid signaling, carbon remobilization, and the production of flavonoids and glutathione could underlie the resistance of Qingliu to leaffolder.

Proteomic analysis of "Oriental Beauty" oolong tea leaves with different degrees of leafhopper infestation.

RATIONALE: Oriental Beauty, a type of oolong tea native to Taiwan, is highly prized by connoisseurs for its unique fruity aroma and sweet taste. Leaves of Oriental Beauty vary in appearance, aroma, and taste, depending on the degree of tea green leafhopper (Jacobiasca formosana) infestation. In this study, the aim is to investigate the differential expression of proteins in leaves with low, medium, and high degrees of leafhopper infestation. METHODS: Proteomic techniques 2DE (two-dimensional electrophoresis) and nanoscale liquid chromatography/tandem mass spectrometry (LC/MS/MS) were used to investigate the differential expression of proteins in tea leaves with different degrees of leafhopper infestation. RESULTS: A total of 89 proteins were found to exhibit significant differences in expression. In a gene ontology analysis, most of these proteins participated in biosynthesis, carbohydrate metabolism, transport, responses to stress, and amino acid metabolism. CONCLUSIONS: These results indicated that the unique aroma and taste of the leaves might be influenced by their protein expression profiles, as well as related factors such as defensive responses to tea green leafhopper saliva.

Preparation and comparison of Fe3O4@graphene oxide nanoclusters for analysis of glimepiride in urine by surface-assisted laser desorption/ionization time-of-flight mass spectrometry.

Graphene oxide (GO) has the ability to absorb certain compounds, and it can be modified with functional groups for different purposes; for instance, iron oxide (IO) nanoparticles can be used to concentrate analyte by a magnet. Recently, many kinds of GO have been developed, such as single-layer GO (SLGO), two-to-four layers of GO (i.e., few-layer GO, FLGO2-4), and four-to-eight layers of GO (i.e., multi-layer GO, MLGO4-8). However, the abilities of these layered GO coated with IO nanoparticles have not been investigated. In this study, we conducted a novel analysis of glimepiride by using layered GO-coated magnetic clusters of IO nanoparticles that were synthesized through a simple and facile emulsion-solvent evaporation method. The methodology is based on (i) enrichment of glimepiride using the layered GO-coated magnetic clusters of IO nanoparticles (IO@SLGO, IO@FLGO2-4, and IO@MLGO4-8), and (ii) rapid determination using magnetic cluster-based surface-assisted laser desorption/ionization time-of-flight mass spectrometry (SALDI-TOFMS). We found that IO@MLGO4-8, the magnetic cluster with the greatest number of GO layers, had the best limit of detection (28.6 pmol/µL for glimepiride). The number of GO layers played a significant role in increasing the sensitivity of the SALDI-MS, indicating that the size of GO in the magnetic clusters contributed to the desorption/ionization efficiency. To the best of our knowledge, this is the first study to enrich glimepiride using magnetic clusters of different GO types and to show that the glimepiride in HLB purified urine adsorbed by magnetic clusters can be analyzed by SALDI-TOFMS.

Rapid determination of isoflavones and other bioactive compounds in soybean using SWATH-MS.

Isoflavones are the major bioactive components in soybeans. Sequential window acquisition of all theoretical fragment ions (SWATH) is a kind of data-independent acquisition (DIA), such that all fragments of each precursor will be preserved in a SWATH-Mass Spectrometry (SWATH-MS) run. In this study, a high-throughput SWATH-MS method for the determination of 12 isoflavones in soybeans was established. Furthermore, amino acids, saponins can be semi-quantitated from the same SWATH-MS data. Combination of targeted quantification and untargeted profiling with SWATH, all bioactive compounds were analyzed within 5 min in 10 min run time, and the method had good linear regression with r2 > 0.99. The precisions (RSD %) of the intra-day and inter-day analyses ranged from 2.11% to 18.7%, and the accuracies (RE%) ranged from -14.39% to 17.48%. The matrix effect ranged from 88.66% to 114.82%. Moreover, 7 varieties of soybeans were analyzed and compared with this robust screening method.

Proteomic analysis of rat kidney under maleic acid treatment by SWATH-MS technology.

RATIONALE: Maleic acid is an industrial-grade chemical that is often used in adhesives, stabilizers, and preservatives. It is unknown whether long-term consumption of maleic acid modified starch is harmful to humans. However, many studies have indicated that maleic acid causes renal tubular damage in animal models, even as the associated pathways remain unclear. Sequential window acquisition of all theoretical fragment ion spectra (SWATH) is the most innovative of the label-free quantitative technologies which have better quantification performance. Therefore, SWATH technology was used to investigate the effect of maleic acid on the rat kidney proteome in this study. METHODS: Sprague-Dawley(SD) rats were treated with 0 mg/kg (control), 6 mg/kg (low-dose), 10 mg/kg (medium-dose), and 60 mg/kg (high-dose) of maleic acid. After kidney protein extraction, 28% SDS-PAGE was used, followed by in-gel digestion and desalting. Next, the samples were analyzed with ultra-performance liquid chromatography (UPLC) coupled with quadrupole time-of-flight mass spectrometry (Q-TOF MS), and data-dependent acquisition (DDA) and SWATH technology were also used. The gene ontology and pathway analysis were accomplished. Ultimately, these protein biomarkers were validated by using scheduled high-resolution multiple reaction monitoring (sMRMHR ). RESULTS: Comparisons of the control group with the other three groups revealed that 95, 130, and 103 proteins were expressed at significantly different levels in the control group and in the low-dose, medium-dose, and high-dose groups, respectively. According to the gene ontology analysis, the major processes that these proteins were involved in were metabolic processes, biological regulation, cellular processes, and responses to stimuli; the major functions that these proteins were involved in were binding, hydrolase activity, catalytic activity, and oxidoreductase activity; and the major cellular components hat they were involved in were the cytoplasm, extracellular region, membrane, and mitochondria. According to the KEGG pathway analysis, these proteins were involved in 35 pathways, five of which, the carbohydrate metabolism, folate biosynthesis, renal tubular resorption, amino acid metabolism, and Ras signaling pathways, are discussed in this study. Ultimately, 19 proteins involved in 12 important pathways were validated by sMRMHR . CONCLUSIONS: It was demonstrated that maleic acid caused insufficient energy production, which might lead to a decrease in the activity of the sodium-potassium ATP pump and hydrogen ion ATP pump, which could in turn have caused renal tubular resorption and hydrogen ion regulation to be blocked, thus leading to the accumulation of hydrogen ions in the renal tubules, which would then result in renal tubular acidification followed finally by Fanconi syndrome.

Rapid determination of bioactive compounds in the different organs of Salvia Miltiorrhiza by UPLC-MS/MS.

Salvia miltiorrhiza has been widely used in Asia for medicinal purposes for >1000 years due to the high levels of bioactive constituents it contains. In this study, a simple and rapid ultrasound-assisted liquid extraction (5 min) was applied for the extraction of these bioactive constituents. The extracts were analyzed by using rapid ultrahigh performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) with simultaneous positive and negative electrospray ionization in a single analytical run. Eight analytes were separated within 2.2 min during 6 min of run time with UHPLC-MS/MS operated in multiple reaction monitoring (MRM) mode. The concentration of salvianolic acids and tanshinones in the different organs of different varieties of Salvia miltiorrhiza ranged from 6.4 to 382.1 mg/g and 0.03 to 31.7 mg/g, respectively. To the best of our knowledge, this is the first report to characterize the tanshinone compounds found in the flower and stem/leaf of Salvia miltiorrhiza by UHPLC-MS/MS.

A rapid, simple, and high-throughput UPLC-MS/MS method for simultaneous determination of bioactive constituents in Salvia miltiorrhiza with positive/negative ionization switching.

The dried root of Danshen (Salvia miltiorrhiza) is a Chinese medicine that has been used in traditional treatments for more than 2000 years in Asia. It has also been used to treat cardiovascular diseases for decades. The goal of this study was to develop a new, fast, and simple ultrahigh-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) analytical method for the analysis of phenolic acids and tanshinones in Danshen. The samples were separated on a Kinetex F5 column (2.1 x 100 mm, 1.7 µm, pentafluorophenyl) using 78% acetonitrile with 0.1% formic acid as the mobile phase with isocratic elution while the flow rate was set at 0.45 mL/min. The analysis was performed in a Waters Xevo TQ mass spectrometer via multiple reaction monitoring (MRM) in polarity switching mode. The linearity values of the calibration curves of eight compounds were in the range from 3 to 3200 ng/mL (r2 > 0.99). Method validation was performed in terms of linearity, the relative standard deviations (RSDs) of the intra-day and inter-day, and it was found that precisions were within 15% and the accuracy ranged from 93% to 118%. The recovery was more than 87%. The limit of detection (LOD) and limit of quantification (LOQ) values of the eight analytes varied between 0.023 to 0.75 ng/mL and 0.375 to 1.5 ng/mL, respectively. This approach has the shortest analysis time for the separation of phenolic acids and tanshinones in Danshen using a UPLC core-shell column with F5 stationary phase. Meanwhile, this robust and high-throughput method not only has a short run time (2 min) but also reduce the consumption of organic solvents.

Comparative Proteomic Analysis of Rat Bronchoalveolar Lavage Fluid after Exposure to Zinc Oxide Nanoparticles.

Zinc oxide nanoparticles (ZnO NPs) are one of the most widely used nanomaterials in consumer products and industrial applications. As a result of all these uses, this has raised concerns regarding their potential toxicity. We previously found that candidate markers of idiopathic pulmonary fibrosis and lung cancer were significantly up-regulated in rat bronchoalveolar lavage fluid (BALF) following exposure to ZnO NPs by using a liquid chromatography (LC)-based proteomic approach. To achieve comprehensive protein identification analysis, we conducted the two-dimensional gel electrophosis (2-DE)-based proteomic workflow to analyze the differences in BALF proteins from rats that had been exposed to a high dose of 35 nm ZnO NPs. A total of 31 differentially expressed protein spots were excised from the gels and analyzed by nanoLC-tandem mass spectrometry (MS/MS). Gene ontology (GO) annotation of these proteins showed that most of the differentially expressed proteins were involved in response to stimulus and inflammatory response processes. Moreover, pulmonary surfactant-associated protein D and gelsolin, biomarkers of idiopathic pulmonary fibrosis, were significantly up-regulated in rat BALF after ZnO NPs exposure (2.42- and 2.84-fold, respectively). The results obtained from this present study could provide a complementary consequence with our previous study and contribute to a better understanding of the molecular mechanisms involved in ZnO NP-induced lung disorders.

Metabolic Disturbances in Adult-Onset Still's Disease Evaluated Using Liquid Chromatography/Mass Spectrometry-Based Metabolomic Analysis.

OBJECTIVE: Liquid chromatography/mass spectrometry (LC/MS)-based comprehensive analysis of metabolic profiles with metabolomics approach has potential diagnostic and predictive implications. However, no metabolomics data have been reported in adult-onset Still's disease (AOSD). This study investigated the metabolomic profiles in AOSD patients and examined their association with clinical characteristics and disease outcome. METHODS: Serum metabolite profiles were determined on 32 AOSD patients and 30 healthy controls (HC) using ultra-performance liquid chromatography (UPLC)/MS analysis, and the differentially expressed metabolites were quantified using multiple reactions monitoring (MRM)/MS analysis in 44 patients and 42 HC. Pure standards were utilized to confirm the presence of the differentially expressed metabolites. RESULTS: Eighteen differentially expressed metabolites were identified in AOSD patents using LC/MS-based analysis, of which 13 metabolites were validated by MRM/MS analysis. Among them, serum levels of lysoPC(18:2), urocanic acid and indole were significantly lower, and L-phenylalanine levels were significantly higher in AOSD patients compared with HC. Moreover, serum levels of lysoPC(18:2), PhePhe, uridine, taurine, L-threonine, and (R)-3-Hydroxy-hexadecanoic acid were significantly correlated with disease activity scores (all p<0.05) in AOSD patients. A different clustering of metabolites was associated with a different disease outcome, with significantly lower levels of isovalerylsarcosine observed in patients with chronic articular pattern (median, 77.0AU/ml) compared with monocyclic (341.5AU/ml, p<0.01) or polycyclic systemic pattern (168.0AU/ml, p<0.05). CONCLUSION: Thirteen differentially expressed metabolites identified and validated in AOSD patients were shown to be involved in five metabolic pathways. Significant associations of metabolic profiles with disease activity and outcome of AOSD suggest their involvement in AOSD pathogenesis.

2-DE combined with two-layer feature selection accurately establishes the origin of oolong tea.

Taiwan is known for its high quality oolong tea. Because of high consumer demand, some tea manufactures mix lower quality leaves with genuine Taiwan oolong tea in order to increase profits. Robust scientific methods are, therefore, needed to verify the origin and quality of tea leaves. In this study, we investigated whether two-dimensional gel electrophoresis (2-DE) and nanoscale liquid chromatography/tandem mass spectroscopy (nano-LC/MS/MS) coupled with a two-layer feature selection mechanism comprising information gain attribute evaluation (IGAE) and support vector machine feature selection (SVM-FS) are useful in identifying characteristic proteins that can be used as markers of the original source of oolong tea. Samples in this study included oolong tea leaves from 23 different sources. We found that our method had an accuracy of 95.5% in correctly identifying the origin of the leaves. Overall, our method is a novel approach for determining the origin of oolong tea leaves.

Graphene flakes enhance the detection of TiO2-enriched catechins by SALDI-MS after microwave-assisted enrichment.

In this study, we describe a novel method for the rapid detection of catechins in tea leaves using surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS). The method involves incubating catechins in a mixture of titanium dioxide (TiO2) nanoparticles and graphene flakes (GF) and then subjecting the mixture to microwave irradiation to facilitate the enrichment of analytes. In this approach, TiO2 nanoparticles serve as probes to enrich catechins while GF serve to increase the desorption/ionization efficiency. The use of the TiO2-GF mixture as matrix provides a number of advantages over TiO2 nanoparticles alone, including higher desorption/ionization efficiency, enhanced analyte enrichment and reduced run time (less 10min). We demonstrate that the addition of GF to nanoparticles of TiO2 improves the detection of catechins by SALDI-MS. At a signal-to-noise ratio of 3, the limits of detection were 0.04pmole for (+)-catechin (C), 0.17pmole for (-)-epigallocatechin (EGC), 0.35pmole for (-)-epicatechin gallate (ECG), and 0.50pmole for (-)-epigallocatechin gallate (EGCG). This method was further applied to determine the catechin content in several tea samples and the results showed good shot-to-shot and sample-to-sample reproducibility (less than 10% and 13%, respectively) as well as good quantitative linearity for the four analytes (R(2)>0.995). The precision calculated as percent relative standard derivation (%RSD), ranged from 3.6% to 12.4%. Overall, we show that this method is a sensitive, simple and high-throughput technique for the detection of catechins in tea.

Changes in protein expression in rat bronchoalveolar lavage fluid after exposure to zinc oxide nanoparticles: an iTRAQ proteomic approach.

RATIONALE: Zinc oxide nanoparticles (ZnO NPs) are widely used in consumer products and various biomedical fields. As a result, humans are frequently exposed to these NPs. However, there is a lack of information about the proteins that are expressed in the airway in response to exposure to ZnO NPs. METHODS: Bronchoalveolar lavage fluid (BALF) from Sprague-Dawley (SD) rats that had been exposed to high-dose 35 nm ZnO NPs (N = 6) and filtered air (N = 4) was collected and then labeled with isobaric tags for relative and absolute quantitation (iTRAQ). The differentially expressed proteins were identified by two-dimensional liquid chromatography/tandem mass spectrometry (2D-LC/MS/MS) and further classified by Gene Ontology (GO) annotation. RESULTS: A total of 46 proteins displayed significant changes after exposure. GO annotation of these differentially expressed proteins indicated that exposure to ZnO NPs mainly affected immune and inflammatory processes. Furthermore, S100A8 and S100A9, candidate markers of idiopathic pulmonary fibrosis and lung cancer, were significantly up-regulated (2.78- and 2.87-fold, respectively) following exposure. CONCLUSIONS: Our data are consistent with recent study results that exposure to ZnO NPs induces lung inflammation. These data contribute to a better understanding of how exposure to ZnO NPs leads to lung damage through the functional classification of these proteins.