Ultra-performance Liquid Chromatography-triple Quadrupole Research Articles

Simultaneous determination of cucurbitacin B, E and I in plasma, urine, and melon and fruit vegetables by ultra performance liquid chromatography-triple quadrupole mass spectrometry using atmospheric pressure chemical ionization

A method for the determination of cucurbitacin B (CuB), cucurbitacin I (CuI) and cucurbitacin E (CuE) in plasma, urine and melon and fruit vegetables was developed by ultra performance liquid chromatography-triple quadrupole mass spectrometry (UPLC-MS/MS). The target analytes in plasma and urine were extracted and cleaned-up by solid supported liquid-liquid extraction, while those in melon and fruit vegetables were extracted with acetonitrile and then diluted with water. CuB, CuI and CuE were separated on an XBridge BEH C18 column (100 mm×3.0 mm, 2.5 μm) with gradient elution using mobile phases of methanol and 0.025% (v/v) ammonia aqueous solution. An atmospheric pressure chemical ionization interface was used as the ion source and the analysis was performed in negative ionization multiple reaction monitoring (MRM) mode. The cucurbitacins in plasma and urine were quantified by the matrix working standard curve internal standard method, while those in melon and fruit vegetables were quantified by the solvent standard curve external standard method. Oleandrin was used as the internal standard. The average recoveries were 89.0%-113% for the three cucurbitacins in plasma and urine, with RSDs of 1.7%-12.2% (n=6). The average recoveries were 87.6%-114% for the three cucurbitacins in melon and fruit vegetables, with RSDs of 4.1%-11.1% (n=6). The limit of detection (S/N=3) of the three cucurbitacins was 0.03 μg/L in plasma and urine, and 5-10 μg/kg in melon and fruit vegetables. The method is simple, sensitive and accurate. It has been used for the determination of cucurbitacins in bitter bottle gourd and in the plasma and urine of patients poisoned by bitter bottle gourd, CuB was successfully detected.

Development and validation of a rapid, robust and sensitive UPLC-QQQ-MS/MS method for simultaneous quantification of GSH metabolism in lung cancer cells

Changes in cellular metabolism accompany tumor therapeutic resistance. Metabolite concentrations specifically reflect the cellular state. Glutathione (GSH) metabolism maintains the redox homeostasis while also confers therapeutic resistance to cancer cells. However, analytical methods for studying GSH metabolism rely on high-resolution-based untargeted metabolomics. Since the aim of untargeted metabolomics studies is covering as much metabolites as possible, these methods lack sensitivity for simultaneous analysis of intracellular GSH-related metabolites with different polarities and structures. In this study, based on cultured lung cancer cells, we described a rapid, robust and sensitive ultra-performance liquid chromatography-triple quadrupole tandem mass spectrographic method (UPLC-QQQ-MS/MS) to simultaneously quantify a repertoire of GSH-related metabolites, including GSH, GSSG, glycine, cysteine, glutamine, glutamate, cystine, γ-glutamyl-cysteine and cysteinyl-glycine. This method avoided the use of derivatization and/or ion-pairing reagents and was validated according to United States Food and Drug Administration (US FDA) criteria. The lower limit of quantification was determined to be 0.5–100 ng/mL with lower limits of detection at 0.14–10.07 ng/mL. The intra- and inter-day precision values for all the analytes were <15% CV, and the accuracy ranged from 85.4% to 114% at three levels of quality control. This method combined simple preparation with rapid analytical procedure (8 min), allowed for high-throughput analysis of GSH metabolism in basic and therapeutic treatment conditions within cultured cells. Our data showed a significant alteration of GSH metabolism in two independent resistant cells compared to sensitive cells. This method monitored the impact of molecularly targeted drugs on GSH metabolism within lung cancer cells and therefore helped identifying potential metabolic vulnerability for the therapeutic resistance in lung cancer.

β-carbolines norharman and harman in vegetable oils in China

ABSTRACT The beta-carbolines norharman and harman, two heterocyclic aromatic amines with potential mutagenicity, have been determined in vegetable oils. Identification and analysis were carried out by ultra-performance liquid chromatography-triple quadrupole tandem mass spectrometry (UPLC-MS/MS). In 88 samples analysed, the concentrations of norharman and harman were < LOD to 336.22 ng/g and < LOD to 505.14 ng/g, respectively. A high variability of norharman and harman levels among different oil types was observed. Sesame-, flaxseed-, sunflower seed-, peanut- and rapeseed oils were most contaminated. Both β-carbolines were most likely formed during roasting of the oilseeds. Oil consumption, especially of oils obtained after roasting of the seeds, was a major dietary source of the β-carbolines norharman and harman. Under existing oil risk factors, this investigation contributes to the unprecedented and essential information for dietary assessments associated with oil consumption.

Determination of seven heterocyclic aromatic amines in oils and fried foods by ultra performance liquid chromatography-triple quadrupole mass spectrometry

An ultra performance liquid chromatography-triple quadrupole mass spectrometry (UPLC-MS/MS) method was developed for the determination of seven heterocyclic aromatic amines (HCAs) in oil and fried food samples. The samples were extracted with acetonitrile containing 1% (v/v) ammonia water, defatted by n-hexane saturated with acetonitrile, and cleaned up with PCX solid phase extraction column. The extracts were separated on a Waters ACQUITY UPLC BEH C18 reversed-phase column (50 mm×2.1 mm, 1.7 μm) with gradient elution by using acetonitrile-10 mmol/L ammonium formate aqueous solution as the mobile phase. Quantitative detection was performed by using a positive-ion electrospray ionization source in the multiple reaction monitoring (MRM) mode, employing an internal isotope standard. The method showed good linear relationships in the range of 0.5-100 μg/L for the seven HCAs with correlation coefficients (R2)>0.999. The average recoveries ranged from 64.31% to 113.8% with the relative standard deviations of 0.18%-9.26% at the three spiked levels in oil and fried food samples. The limits of detection (LODs) and limits of quantification (LOQs) were 0.01-0.14 ng/g and 0.09-0.38 ng/g, respectively. The method is sensitive, accurate, and suitable for the determination and confirmation of the seven HCAs in oil and fried food samples.

Defensive Responses of Tea Plants (Camellia sinensis) Against Tea Green Leafhopper Attack: A Multi-Omics Study.

Tea green leafhopper [Empoasca (Matsumurasca) onukii Matsuda] is one of the most devastating pests of tea plants (Camellia sinensis), greatly impacting tea yield and quality. A thorough understanding of the interactions between the tea green leafhopper and the tea plant would facilitate a better pest management. To gain more insights into the molecular and biochemical mechanisms behind their interactions, a combined analysis of the global transcriptome and metabolome reconfiguration of the tea plant challenged with tea green leafhoppers was performed for the first time, complemented with phytohormone analysis. Non-targeted metabolomics analysis by ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-QTOF MS), together with quantifications by ultra-performance liquid chromatography triple quadrupole mass spectrometry (UPLC-QqQ MS), revealed a marked accumulation of various flavonoid compounds and glycosidically bound volatiles but a great reduction in the level of amino acids and glutathione upon leaf herbivory. RNA-Seq data analysis showed a clear modulation of processes related to plant defense. Genes pertaining to the biosynthesis of phenylpropanoids and flavonoids, plant-pathogen interactions, and the biosynthesis of cuticle wax were significantly up-regulated. In particular, the transcript level for a CER1 homolog involved in cuticular wax alkane formation was most drastically elevated and an increase in C29 alkane levels in tea leaf waxes was observed. The tea green leafhopper attack triggered a significant increase in salicylic acid (SA) and a minor increase in jasmonic acid (JA) in infested tea leaves. Moreover, transcription factors (TFs) constitute a large portion of differentially expressed genes, with several TFs families likely involved in SA and JA signaling being significantly induced by tea green leafhopper feeding. This study presents a valuable resource for uncovering insect-induced genes and metabolites, which can potentially be used to enhance insect resistance in tea plants.

Facile in-situ polymerization of polyaniline-functionalized melamine sponge preparation for mass spectrometric monitoring of perfluorooctanoic acid and perfluorooctane sulfonate from biological samples

In this present work, a novel polyaniline-functionalized melamine sponge (PMs) was successfully prepared using a simple unstirred in-situ polymerization process. The PMs was characterized using a scanning electron microscope and contact angle measurements. Its adsorption performance was initially determined via dye adsorption assays, and the conditions affecting the synthesis including polymerization time, acidity, molar ratio, and number and sizes of raw melamine sponge were optimized. The PMs was then used as an efficient adsorbent for the development of a novel, low-cost method for the detection of two representative perfluorinated chemicals, perfluorooctanoic acid (PFOA) and perfluorooctane sulfonate (PFOS), using ultra-performance liquid chromatography-triple quadrupole mass spectrometry (UPLC-QqQ-MS/MS) with the internal standard method. To achieve the best extraction efficiency with this method, several variables were optimized, including adsorption time, pH value, the number of PMs, and desorption conditions. Calibration graphs showed a good linear degree at concentration ranging from 0.1 to 50 μg L−1 for PFOA and 0.01–10 μg L−1 for PFOS, with a coefficient of detection R2 = 0.998. The intra-day and inter-day relative standard deviations were found to range from 5.9% to 8.2% for PFOA, and 5.5% to 7.7% for PFOS. Under these optimized conditions, the method was successfully used to measure PFOA and PFOS content in real human serum and urine samples, with average spiked recoveries ranging from 79% and 91% for PFOA, and 81% to 87% for PFOS.

Occurrence of 40 pharmaceutically active compounds in hospital and urban wastewaters and their contribution to Mahdia coastal seawater contamination.

In the present study, the occurrence of 40 pharmaceuticals belonging to several therapeutic groups was investigated for the first time in hospital effluent, wastewater treatment plant influent and effluent, and seawater in Mahdia, Tunisia. Forty-six samples were collected within a 6-month sampling period. Pharmaceuticals were analyzed using solid-phase extraction followed by ultra-performance liquid chromatography-triple quadrupole mass spectrometry. Thirty-three out of the forty target compounds were detected over a wide concentration of ranges, from nanograms per liter to micrograms per liter, depending on the type of sample. Maximum values were detected for caffeine at 902 μgL-1 in hospital wastewater. This compound, as well as salicylic acid, sulfadiazine, and sulfamethizole, were detected in all samples. The average concentration of total pharmaceuticals in hospital wastewater (340 μgL-1) was higher than those detected in influent and effluent wastewater and seawater (275.11 and 0.2 μgL-1, respectively). Risk quotients (RQs) were also estimated to provide a preliminary environmental risk assessment and results revealed that sulfadiazine, sulfamethoxazole, and fluoxetine could pose medium/high risk to the tested aquatic organisms for maximum measured concentrations in wastewater (including hospital and WWTP samples). Although the measured environmental concentrations (MECs) detected in seawater samples might not pose a toxic effect to the aquatic organisms (except for salicylic acid, sulfamethoxazole and fluoxetine), further researches are needed due to the continuous release of wastewater in the environment and the limited efficiency of wastewater treatment processes.

Monitoring DNA adducts in human blood samples using magnetic Fe3O4@graphene oxide as a nano-adsorbent and mass spectrometry

DNA adducts usually occur at a level of 0.1–1 adducts per 108 unmodified DNA bases, which can cause many adverse health effects if not normally repaired. Using microgram amounts of DNA to detect DNA adducts remains challenging due to the extremely low levels. Commercial solid phase extraction is the most widely used method for extracting DNA adducts from biological samples, but it is time consuming and costly, and the hydrophobic interactions between various alkyl-bonded phases and analytes typically lack selectivity when used to extract DNA adducts. Therefore, an effective method for DNA adducts analysis based on Fe3O4@graphene oxide (GO) nano-adsorbent and ultra-performance liquid chromatography-triple quadrupole mass spectrometry detection was developed in this study, in which the selectivity may also be increased by π-π stacking interactions between GO and adduct molecules. Several variables, including the extraction time, extraction pH, nano-sorbent amount and elution solvent were optimized, to achieve the best extraction efficiency using the proposed method. A small amount of Fe3O4@GO (2.5 mg) exhibited a good adsorption performance for the model DNA adducts, and butanol containing 4% NH3·H2O showed good elution effects. Under the optimized conditions, satisfying recoveries (78–107%) from calf thymus DNA samples were achieved, and 2 adducts were detected in only 2 μg of blood DNA. This method can be used as an effective strategy for DNA adduct analysis, and can be extended to other biological samples.

Application of ultra performance liquid chromatography- triple quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS) for the determination of antibiotic residues in meat and fish

The residue of antibiotics in meat and meat products is one of the social problems for its effects and risks to human health, especially in the antibiotic resistance crisis. Nowadays, there are hundreds of drugs commercially available, with difference biochemical characteristics. Therefore, it is necessary to determine and monitor these residues in meat and meat products. The methods using liquid chromatography (LC) and ELISA are widely applied in numbers of laboratories to determine specified substances. In several laboratories, the state-of-the-art highresolution analytical technique, using Ultra Performance Liquid chromatography-triple quadrupole Time-of-flight mass spectrometry (UPLC-QTOF-MS), is applied to analyse the knowns and unknowns residues in food. In this study, we used this technique for screening the residue of antibiotics, banned substances (Sulfonamids, Quinolones, Tetracyclines, &amp;beta;-Agonists&amp;hellip;) and other toxins in pork, chicken, beef and fish samples. Ninety-three randomly collected samples have been screened and residues of Amoxicillin, Cefalexin and Ampicillin have been detected in three of them.

Determination of Flavonoid Glycosides by UPLC-MS to Authenticate Commercial Lemonade.

So far, there is no report on the quality evaluation of lemonade available in the market. In this study, a sample preparation method was developed for the determination of flavonoid glycosides by ultra-performance liquid chromatography–mass spectrometry (UPLC-MS) based on vortex-assisted dispersive liquid-liquid microextraction. First, potential flavonoids in lemonade were scanned and identified by ultra-performance liquid chromatography–time of flight mass spectrometry (UPLC-TOF/MS). Five flavonoid glycosides were identified as eriocitrin, narirutin, hesperidin, rutin, and diosmin according to the molecular formula provided by TOF/MS and subsequent confirmation of the authentic standard. Then, an ultra-performance liquid chromatography–triple quadrupole mass spectrometry (UPLC-QqQ/MS) method was developed to determine these five flavonoid glycosides in lemonade. The results showed that the content of rutin in some lemonade was unreasonably high. We suspected that many illegal manufacturers achieved the goal of low-cost counterfeiting lemonade by adding rutin. This suggested that it was necessary for relevant departments of the state to make stricter regulations on the quality standards of lemonade beverages.

Optimizing Ultrasound-Assisted Deep Eutectic Solvent Extraction of Bioactive Compounds from Chinese Wild Rice.

In this study, deep eutectic solvents (DESs) were used for the ultrasound-assisted extraction (UAE) of valuable bioactive compounds from Chinese wild rice (Zizania spp.). To this end, 7 different choline chloride (CC)-based DESs were tested as green extraction solvents. Choline chloride/1,4-butanediol (DES-2) exhibited the best extraction efficiency in terms of parameters such as the total flavonoid content (TFC), total phenolic content (TPC), and free radical scavenging capacity (DPPH● and ABTS●+). Subsequently, the UAE procedure using 76.6% DES-2 was also optimized: An extraction temperature of 51.2 °C and a solid–liquid ratio of 37.0 mg/mL were considered optimal by a Box–Behnken experiment. The optimized extraction procedure proved efficient for the extraction of 9 phenolic and 3 flavonoid compounds from Chinese wild rice as determined by quantification based on ultra-performance liquid chromatography–triple quadrupole tandem mass spectrometry (UPLC-QqQ-MS). This work, thus, demonstrates the possibility of customizing green solvents that offer greater extraction capacity than that of organic solvents.

Comparative quality of the forms of decoction pieces evaluated by multidimensional chemical analysis and chemometrics: Poria cocos, a pilot study

Many Chinese medicinal materials (CMMs) are parts of plants or fungi that have been processed into different physical forms, termed decoction pieces, that are typically boiled in water for consumption. One CMM may have several decoction pieces forms, e.g., slices, small cubes (dice), or grains. The specifications that have different morphological parameters (shape, size and thickness) for these various decoction pieces have been developed over, in some cases, centuries of practice. Nevertheless, whether and how the form of decoction pieces affects the extraction (decoction) dynamics, and quality stability during storage has not been studied. Here, we investigated Poria cocos (PC) as a pilot study; we explore how the form of PC decoction pieces affects its chemistry using multidimensional chemical evaluation such as ultra-performance liquid chromatography-photodiode array-quadrupole time-of-flight mass spectrometry (UHPLC-PDA-QTOF-MS/MS), ultra-performance liquid chromatography-triple quadrupole mass spectrometry (UHPLC-QqQ-MS/MS) and high performance gel permeation chromatography coupled with charged aerosol detector (HPGPC-CAD), combined with analysis of variance (ANOVA), principal component analysis (PCA), factor analysis (FA) and hierarchical cluster analysis (HCA). The results indicated that different specifications had significant differences, and these specifications could be divided into four groups. The comprehensive results of the chemical analyses undertaken here indicate that the highest potentially available quality of PC decoction pieces was in the forms of curl, ultra-small grains and small grains, followed by thin slices. This information not only is conducive to promoting the standardization of the specification/form of PC decoction pieces and maximizing the benefits from its utilization, but also provide a promising strategy for assessing other CMM decoction pieces in different forms.

Simultaneous determination of seven effective components of Tripterygium glycosides in human biological matrices by ultra performance liquid chromatography–triple quadrupole mass spectrometry

This paper developed a novel, sensitive, and selective ultra-performance liquid chromatography–triple quad mass spectrometry method to simultaneously determine seven effective constituents (triptolide, triptophenolide, celastrol, wilforgine, wilforine, wilfordine and wilfortrine) of Tripterygium glycosides (GTW) in human serum and urine. The chromatographic separation was performed on the C18 column using an ammonium acetate buffer solution–acetonitrile (both containing 0.1% formic acid) in a gradient program with a flow rate of 0.3 mL/min. Monitoring reaction mode was applied to target compounds quantitative analysis in the positive electrospray ionization (ESI) mode. The analysis process took 11 min in total. This method was fully validated with a linear range of 1–200 ng/mL for triptolide, 0.4–80 ng/mL for celastrol, 0.1–20 ng/mL for triptophenolide, wilforgine, wilforine, wilfordine, and wilfortrine. The intra-day and inter-day accuracy and precision of the target compounds all met the 15% criterion in both serum and urine. Extraction recovery, matrix effect, and dilution integrity were also validated. The short-term and long-term stability results indicated that all the constituents were stable in human serum and urine under the investigated storage conditions. 10 patients' specimens were collected and analyzed. Most of the compounds exhibited the tendency of higher concentration in urine than that in serum. The concentration that was detected in the serum and in the urine of alkaloids showed a positive-correlation property. This is the first time that triptophenolide was quantified in human bio-matrices. The method is feasible for multi-components therapeutic monitoring or pharmacokinetics study in clinical pharmaceutical research of Tripterygium glycosides.

Effects of nitrogen supply on flavonol glycoside biosynthesis and accumulation in tea leaves (Camellia sinensis)

Widely distributed in tea plants, the flavonoid flavonol and its glycosylated derivatives have important roles in determining tea quality. However, the biosynthesis and accumulation of these compounds has not been fully studied, especially in response to nitrogen (N) supply. In the present study, ‘Longjing 43’ potted tea seedlings were subjected to N deficiency (0g/pot), normal N (4g/pot) or excess N (16g/pot). Quantitative analyses using Ultra Performance Liquid Chromatography-Triple Quadrupole Mass Spectrometry (UPLC-QqQ-MS/MS) revealed that most flavonol glycosides (e.g., Quercetin-3-glucoside, Kaempferol-3-rgalactoside and Kaempferol-3-glucosyl-rhamnsoyl-glucoside) accumulated to the highest levels when treated with normal N. Results from metabolomics using Gas Chromatography-Mass Spectrometer (GC-MS) suggested that the levels of carbohydrate substrates of flavonol glycosides (e.g., sucrose, sucrose-6-phosphate, D-fructose 1,6-bisphosphate and glucose-1-phosphate) were positively correlated with flavonol glycoside content in response to N availability. Furthermore, Quantitative Real-time PCR analysis of 28 genes confirmed that genes related to flavonoid (e.g., flavonol synthase 1, flavonol 3-O-galactosyltransferase) and carbohydrate (e.g., sucrose phosphate synthase, sucrose synthase and glucokinase) metabolism have important roles in regulating the biosynthesis and accumulation of flavonol glycosides. Collectively, our results suggest that normal N levels promote the biosynthesis of flavonol glycosides through gene regulation and the accumulation of substrate carbohydrates, while abnormal N availability has inhibitory effects, especially excess N.

Determination of coriatin and corianin in plasma and urine using ultra-performance liquid chromatography-triple quadrupole mass spectrometry

An ultra-performance liquid chromatography-triple quadrupole mass spectrometry (UPLC-MS/MS) method has been developed for the determination of coriatin and corianin in plasma and urine, which are the biomarkers of poisoning caused by Coriaria sinica Maxim. Plasma and urine samples were extracted and purified using a solid supported liquid/liquid extraction method. Chromatographic separation was performed on a Cortecs C18 column (100 mm×2.1 mm, 1.6 μm) using a gradient elution of methanol and water. Coriatin and corianin were detected using negative electrospray ionization tandem mass spectrometry in multiple reaction monitoring (MRM) mode and quantified via a matrix working standard curve internal standard method; florfenicol was used as the internal standard. The assay was linear in the calibration range of 0.03-5.0 μg/L for coriatin and 0.3-50 μg/L for corianin in plasma, and 0.1-10 μg/L and 1-100 μg/L for coriatin and corianin in urine, respectively. The average recoveries were 86.2%-110% for coriatin and corianin in plasma and urine with relative standard deviations of 5.1%-14.6% (n=6). The limits of detection (S/N=3) for coriatin and corianin were 0.01 μg/L and 0.1 μg/L in plasma, and 0.03 μg/L and 0.3 μg/L in urine, respectively. The method is simple, sensitive and accurate for the determination of coriatin and corianin in plasma and urine for toxicological purposes.

Optimization of Vortex-Assisted Dispersive Liquid-Liquid Microextraction for the Simultaneous Quantitation of Eleven Non-Anthocyanin Polyphenols in Commercial Blueberry Using the Multi-Objective Response Surface Methodology and Desirability Function Approach.

In the present study, 11 non-anthocyanin polyphenols, gallic acid, protocatechuate, vanillic acid, syringic acid, ferulic acid, quercetin, catechin, epicatechin, epigallocatechin gallate, gallocatechin gallate and epicatechin gallate—were firstly screened and identified from blueberries using an ultra performance liquid chromatography–time of flight mass spectrography (UPLC-TOF/MS) method. Then, a sample preparation method was developed based on vortex-assisted dispersive liquid-liquid microextraction. The microextraction conditions, including the amount of ethyl acetate, the amount of acetonitrile and the solution pH, were optimized through the multi-objective response surface methodology and desirability function approach. Finally, an ultra performance liquid chromatography–triple quadrupole mass spectrography (UPLC-QqQ/MS) method was developed to determine the 11 non-anthocyanin polyphenols in 25 commercial blueberry samples from Sichuan province and Chongqing city. The results show that this new method with high accuracy, good precision and simple operation characteristics, can be used to determine non-anthocyanin polyphenols in blueberries and is expected to be used in the analysis of other fruits and vegetables.

Toxicokinetics and Metabolism of 3-Monochloropropane 1,2-Diol Dipalmitate in Sprague Dawley Rats.

Fatty acid esters of 3-monochloropropane 1,2-diol (3-MCPD) are a group of processing-induced toxicants. To better clarify their possible toxicological effects and mechanisms, it is important to investigate their absorption, distribution, metabolism, and excretion. In this study, the kinetic parameters of 3-MCPD dipalmitate in Sprague Dawley (SD)rat plasma were determined using ultraperformance liquid chromatography-triplequadrupole mass spectrometry. 3-MCPD dipalmitate was absorbed in rats with a Cmax of 135.00 ng/mL, a T1/2 of 3.87 h, a Tmax of 2.5 h, an MRT of 5.08 h, a CL of 3.50 L/h/g, a Vd of 21.34 L/g, and an AUC0-∞ of 458.47 h·ng/mL. A total of 17 metabolites were identified, and 16 of them were reported for the first time. Furthermore, these metabolites were examined for their presences in the liver, kidney, testis, brain, spleen, thymus, intestine, plasma, feces, and urine samples 2, 6, 12, 24, and 48 h after oral administration of 3-MCPD dipalmitate using Metabolynx software.

CYP3A Activation and Glutathione Depletion Aggravate Emodin-Induced Liver Injury.

1,3,8-Trihydroxy-6-methylanthraquinone (emodin), a widely existing natural product in herbal medicines, has been reported to be hepatotoxic, but the exact underlying mechanism is still not fully understood. The objective of the present study was to evaluate the role of CYP3A and glutathione (GSH) in emodin-induced liver injury. Primary human hepatocytes were exposed to emodin with and without addition of CYP3A inducer/inhibitor and GSH synthesis inhibitor. It was found that emodin-mediated cytotoxicity increased when CYP3A was activated and GSH was depleted. Hepatotoxicity induced by emodin in rats by activation/inhibition of CYP3A and depletion of GSH was further investigated. Administration of emodin in combination with l-buthionine sulfoximine (BSO) or dexamethasone (DEX) resulted in aggravated liver injury, whereas pretreatment with ketoconazole (KTZ) suppressed the side effects caused by emodin. In addition, plasma exposure of emodin and its glucuronide metabolite were measured by ultraperformance liquid chromatography triple quadrupole mass spectrometry. Emodin and its glucuronide were lower in BSO-, DEX-, and KTZ- co-treated rats compared with those administered with emodin alone. In conclusion, these mentioned results suggested that CYP3A induction and GSH depletion might be involved in hepatotoxicity induced by emodin. This study may help to understand the risk factors and the mechanism of hepatotoxicity of emodin in humans.

Effects of vitro sucrose on quality components of tea plants (Camellia sinensis) based on transcriptomic and metabolic analysis

BackgroundTea plants [Camellia sinensis (L.) O. Kuntze] can produce one of the three most widely popular non-alcoholic beverages throughout the world. Polyphenols and volatiles are the main functional ingredients determining tea’s quality and flavor; however, the biotic or abiotic factors affecting tea polyphenol biosynthesis are unclear. This paper focuses on the molecular mechanisms of sucrose on polyphenol biosynthesis and volatile composition variation in tea plants.ResultsMetabolic analysis showed that the total content of anthocyanins, catechins, and proanthocyanidins(PAs) increased with sucrose, and they accumulated most significantly after 14 days of treatment. Transcriptomic analysis revealed 8384 and 5571 differentially expressed genes in 2-day and 14-day sucrose-treated tea plants compared with control-treated plants. Most of the structural genes and transcription factors (TFs) involved in polyphenol biosynthesis were significantly up-regulated after 2d. Among these transcripts, the predicted genes encoding glutathione S-transferase (GST), ATP-binding cassette transporters (ABC transporters), and multidrug and toxic compound extrusion transporters (MATE transporters) appeared up regulated. Correspondingly, ultra-performance liquid chromatography-triple quadrupole mass spectrometry (UPLC-QQQ-MS/MS) analysis revealed that the content of non-galloylated catechins and oligomeric PAs decreased in the upper-stem and increased in the lower-stem significantly, especially catechin (C), epicatechin (EC), and their oligomeric PAs. This result suggests that the related flavonoids were transported downward in the stem by transporters. GC/MS data implied that four types of volatile compounds, namely terpene derivatives, aromatic derivatives, lipid derivatives, and others, were accumulated differently after in vitro sucrose treatment.ConclusionsOur data demonstrated that sucrose regulates polyphenol biosynthesis in Camellia sinensis by altering the expression of transcription factor genes and pathway genes. Additionally, sucrose promotes the transport of polyphenols and changes the aroma composition in tea plant.

A simple multi-residue method for determination of plant growth retardants in Ophiopogon japonicus and soil using ultra-performance liquid chromatography–tandem mass spectrometry

Plant growth retardants may play an important role in regulation of yield and quality of crops, fruits, and vegetables. Such compounds have begun to be used in the cultivation of traditional Chinese medicines (TCMs), especially for root medicines. Although the potential risks to human health of these compounds has attracted increasing attention, analytical methods for detection of plant growth retardants in TCMs remain poorly investigated. In this study, an effective and reliable method for simultaneous determination of 11 plant growth retardants in Ophiopogon japonicus and soil samples was developed by ultra-performance liquid chromatography-triple quadrupole tandem mass spectrometry (UPLC-QqQ-MS/MS). Extraction was conducted in acetonitrile containing 1.0% (v/v) acetic acid with ultrasonication. The octadecylsilyl (C18) and MgSO4 were used as the dispersive-solid phase extraction (d-SPE) sorbent and provided satisfactory recoveries for the analytes. The conditions of extraction and LC-MS/MS were optimized to achieve the highest recovery and sensitivity. Good linearity was achieved within a wide range with all correlation coefficients exceeding 0.9950. The recoveries of all analytes in O. japonicus and soil samples ranged from 57.37% (choline chloride) to 99.93% (trinexapac-ethyl) and from 54.37% (daminozide) to 94.82% (triadimenol), respectively. The limits of quantifications ranged from 0.03 to 3.54 μg/L. The proposed method was successfully applied to detect and quantify 11 plant growth retardants in empirical O. japonicus and soil samples. High frequency of paclobutrazol and choline chloride was found in O. japonicus samples. In addition, paclobutrazol showed a high residual concentration (>1100 μg/kg) in the soil of O. japonicus indigenous production.