Liquid Chromatography-triple Quadrupole Mass Research Articles

Development and validation of an open screening method for doping substances in urine by gas chromatography quadrupole time-of-flight mass spectrometry

In anti-doping, a high number of classes of substances are prohibited and laboratories need to detect these at low urinary concentrations. Traditionally, testing is done using complimentary liquid chromatography mass spectrometry and gas chromatography mass spectrometry. High resolution mass spectrometric acquisition has some important advantages over triple quadrupole instruments (e.g., open screening due to full scan high resolution data acquisition with retrospectivity, compatibility with libraries and a straightforward and effortless addition and validation of new compounds in the future). Doping samples can be stored for 10 years and retrospective data analysis can be used to re-evaluate previously acquired data (e.g., searching for prohibited (designer) substances that were unknown at the initial moment of analysis). During the past decade, these advantages have led to the wide-scale transfer of liquid chromatography triple quadrupole mass spectrometry screening to liquid chromatography high resolution mass spectrometry screening for doping control purposes. Up to now, for gas chromatography a similar switch to high resolution screening has not yet occurred, because so far no method has been developed that combines sufficient sensitivity with wide-scale drug detection.In this work, the current gas chromatography triple quadrupole mass spectrometry screening method for human doping control purposes was successfully converted into an equivalent and complete gas chromatography high resolution acquisition screening method. This new screening method on a gas chromatography quadrupole time-of-flight mass spectrometer has been developed and validated. The method is compliant with the World Anti-Doping Agency requirements and allows the detection of 294 target compounds (and 14 internal standards), including diuretics, stimulants, narcotics, beta-2-agonists, beta-blockers, hormone modulators, anabolic agents and the quantification of 14 endogenous steroids in a single fast run (14.1 min).

Regional differences of hypothermia on oxidative stress following hypoxia-ischemia: a study of DHA and hypothermia on brain lipid peroxidation in newborn piglets.

Background Oxidative stress plays an important part in the pathophysiology of hypoxic-ischemic encephalopathy (HIE) and is reliably measured through prostanoids following lipid peroxidation of polyunsaturated fatty acids (PUFAs). The aim of the study is to measure oxidative stress in the prefrontal cortex, white matter and hippocampus in the brains of hypoxic-ischemic piglets treated with docosahexaenoic acid (DHA) and therapeutic hypothermia (TH) and investigate the additive effects of DHA on hypothermia by factorial design. Methods Fifty-five piglets were randomized as having severe global hypoxia (n=48) or not (sham, n=7). Hypoxic piglets were further randomized: vehicle (VEH), DHA, VEH+hypothermia (HT) or HT+DHA. A total of 5 mg/kg DHA was given intravenously 210 min after the end of hypoxia. Brain tissues were analyzed using liquid chromatography triple quadrupole mass spectrometry technique (LC-MS). A two-way analysis of variance (ANOVA) was performed with DHA and HT as main effects. Results In the white matter, we found main effects of DHA on DH-isoprostanes (P=0.030) and a main effect of HT on F4-neuroprostanes (F4-NeuroPs) (P=0.007), F2-isoprostanes (F2-IsoPs) (P=0.043) and DH-isoprostanes (P=0.023). In the cortex, the ANOVA analysis showed the interactions of main effects between DHA and HT for neurofuranes (NeuroFs) (P=0.092) and DH-isoprostanes (P=0.015) as DHA significantly reduced lipid peroxidation in the absence of HT. DHA compared to VEH significantly reduced NeuroFs (P=0.019) and DH-isoprostanes (P=0.010). No differences were found in the hippocampus. Conclusion After severe hypoxia, HT reduced lipid peroxidation in the white matter but not in the cortical gray matter. HT attenuated the reducing effect of DHA on lipid peroxidation in the cortex. Further studies are needed to determine whether DHA can be an effective add-on therapy for TH.

Rapid quantitative analysis of methylphenidate and ritalinic acid in oral fluid by liquid chromatography triple quadrupole mass spectrometry (LC-QqQ-MS)

Methylphenidate (MPH), which is metabolized into ritalinic acid (RA), is an amphetamine derivative largely used in the treatment of attention-deficit hyperactivity disorder, a neurological condition commonly diagnosed in early childhood. Ensuring that patients comply with clinical treatment is crucial and compliance is generally monitored in blood or urine specimens which, especially in the case of children, can be challenging to obtain on a repetitive basis. Here we report validation of a specific, non-invasive, and rapid dilute-and-shoot analytical method for the detection and quantitation of MPH and RA in oral fluid (OF). The method is based on liquid chromatography coupled to triple quadrupole MS with electrospray ionization utilizing dynamic MRM mode. Subject OF specimens were collected using a Quantisal™ device, processed, and diluted for analysis with seven-point quadratic calibration curves (weighting of 1/x) using MPH-d9 and (±)-threo-RA-d10 as internal standards. QC samples and diluted specimens showed intra- and inter-day bias and imprecision values no greater than ±12%. The LOD and LOQ for MPH were 0.1 and 0.5 ng/mL, respectively, and 0.2 ng/mL and 0.5 ng/mL for RA, respectively, indicating the validity of the method for identification and confirmation at low concentrations. Selectivity was specific for the analytes of interest and matrix effects were minimized through the use of internal standard based quantitation.

PEGylation of Carbonate Apatite Nanoparticles Prevents Opsonin Binding and Enhances Tumor Accumulation of Gemcitabine

pH sensitives carbonate apatite (CA) has emerged as a targeted delivery vehicle for chemotherapeutics agent with tremendous potential to increase the effectivity of breast cancer treatment. The major challenge for intravenous delivery of drug-incorporated nanoparticles is their rapid opsonization, resulting in accumulation within the organs of reticuloendothelial system, such as liver and spleen. Therefore, surface modification by polyethylene glycol was implemented to improve the half-life of drug-particle complexes and enhance their uptake by target tissues. A simple, rapid, and sensitive triple quadrupole liquid chromatography–mass spectrometry method was developed and validated for quantification of gemcitabine in plasma, various organs and tumor tissues of mice with breast carcinoma, whereas sodium dodecyl sulfate-polyacrylamide gel electrophoresis, quadrupole-time of flight liquid chromatography–mass spectrometry and analysis by SwissProt.Mus_musculus database were performed for protein separation, identification, and homology search by comparing the de novo sequence tag. PEGylated CA exhibited almost 6-fold increase in gemcitabine accumulation in tumor with significant reduction in other organs within 1 h of intravenous administration, compared to free drug. In addition, plasma drug amount was found to be higher in PEGylated particles, implying their role in prolonging blood circulation time of particle-bound gemcitabine. Investigation of protein corona composition demonstrated notable reduction in opsonin interactions after PEGylation of CA particles. Overall, the results indicate that the composition and dynamics of protein corona subjected to alteration by PEGylation play crucial roles in affecting successful nanoparticle-based targeted delivery of a cytotoxic drug.

Detection of prohibited substances in equine hair by ultra-high performance liquid chromatography-triple quadrupole mass spectrometry - application to doping control samples.

The detection of drugs in human hair samples has been performed by laboratories around the world for many years and the matrix is popular in disciplines, such as workplace drug testing. To date, however, hair has not become a routinely utilised matrix in sports drug detection. The analysis of hair samples offers several potential advantages to doping control laboratories, not least of which are the greatly extended detection window and the ease of sample collection and storage. This article describes the development, validation, and utilisation of a sensitive ultra-high performance liquid chromatography-triple quadrupole mass spectrometry (UHPLC-MS/MS) method for the detection of 50 compounds. This provides significantly improved coverage for those analytes which would be of particular interest if detected in hair, such as anabolic steroid esters and selective androgen receptor modulators (SARMs). Qualitative validation of the method resulted in estimated limits of detection as low as 0.1pg/mg for the majority of compounds, with all being detected at 2pg/mg or below. The suitability of the method for the detection of prohibited substances in incurred material was demonstrated by the successful detection of several compounds, such as stanozolol, boldenone undecylenate, clenbuterol, and GW-501516, in genuine equine hair samples. Estimated concentrations of the detected substances ranged from 0.27 to 8.6pg/mg. The method has been shown to be fit-for-purpose for routine screening of equine hair samples by the analysis of over 400 genuine hair samples.

Urinary Polyamine Biomarker Panels with Machine-Learning Differentiated Colorectal Cancers, Benign Disease, and Healthy Controls.

Colorectal cancer (CRC) is one of the most daunting diseases due to its increasing worldwide prevalence, which requires imperative development of minimally or non-invasive screening tests. Urinary polyamines have been reported as potential markers to detect CRC, and an accurate pattern recognition to differentiate CRC with early stage cases from healthy controls are needed. Here, we utilized liquid chromatography triple quadrupole mass spectrometry to profile seven kinds of polyamines, such as spermine and spermidine with their acetylated forms. Urinary samples from 201 CRCs and 31 non-CRCs revealed the N1,N12-diacetylspermine showing the highest area under the receiver operating characteristic curve (AUC), 0.794 (the 95% confidence interval (CI): 0.704–0.885, p < 0.0001), to differentiate CRC from the benign and healthy controls. Overall, 59 samples were analyzed to evaluate the reproducibility of quantified concentrations, acquired by collecting three times on three days each from each healthy control. We confirmed the stability of the observed quantified values. A machine learning method using combinations of polyamines showed a higher AUC value of 0.961 (95% CI: 0.937–0.984, p < 0.0001). Computational validations confirmed the generalization ability of the models. Taken together, polyamines and a machine-learning method showed potential as a screening tool of CRC.

Natural variation of hormone levels in Arabidopsis roots and correlations with complex root architecture.

Studies on natural variation are an important tool to unravel the genetic basis of quantitative traits in plants. Despite the significant roles of phytohormones in plant development, including root architecture, hardly any studies have been done to investigate natural variation in endogenous hormone levels in plants. Therefore, in the present study a range of hormones were quantified in root extracts of thirteen Arabidopsis thaliana accessions using a ultra performance liquid chromatography triple quadrupole mass spectrometer. Root system architecture of the set of accessions was quantified, using a new parameter (mature root unit) for complex root systems, and correlated with the phytohormone data. Significant variations in phytohormone levels among the accessions were detected, but were remarkably small, namely less than three‐fold difference between extremes. For cytokinins, relatively larger variations were found for ribosides and glucosides, as compared to the free bases. For root phenotyping, length‐related traits—lateral root length and total root length—showed larger variations than lateral root number‐related ones. For root architecture, antagonistic interactions between hormones, for example, indole‐3‐acetic acid to trans‐zeatin were detected in correlation analysis. These findings provide conclusive evidence for the presence of natural variation in phytohormone levels in Arabidopsis roots, suggesting that quantitative genetic analyses are feasible.

Determination of the low-abundant protein biomarker hCG from dried matrix spots using immunocapture and nano liquid chromatography mass spectrometry

Using LC-MS/MS for determination of low-abundance protein biomarkers from dried blood spots is challenging due to the combination of low biomarker levels (low pM-level) and small sample volumes (typically <50 μL). In the present paper it is demonstrated that use of state-of-the-art nano liquid chromatography triple quadrupole mass spectrometry in combination with immunoaffinity sample clean-up enable determination of the low abundance biomarker human chorionic gonadotropin (hCG) from four different biological matrices (whole blood, serum, plasma and urine) at its upper reference level (low pM). Detection limits for hCG was determined for all matrices from both commercially available non-soluble DBS sampling material (DMPK-C) and the water-soluble material carboxymethyl cellulose (CMC). The detection limits (S/N = 3) were ranging from 5.0 IU/L (14.5 pM; whole blood) to 10.5 IU/L (30.5 pM; urine) for DMPK-C and from 2.1 IU/L (6.1 pM; urine) to 6.4 IU/L (18.6 pM; plasma) for CMC. A brief evaluation was performed for both sampling materials using serum as matrix resulting in sufficient linearity (r2 ≥ 0.93, range 20–1000 IU/mL (58–2900 pM) for DMPK-C and 10–1000 IU/mL (29–2900 pM) for CMC), repeatability (RSD% = 13–31%) and accuracy (95–106%). To demonstrate the applicability of the method to real samples, a serum sample from a patient previously diagnosed with cancer was also analyzed using both sampling materials. The concentration levels found using the two materials were similar (5280± 595 IU/L (15,312 ± 1726 pM, n = 3) in the DMPK-C spot and 5060 ± 430 IU/L (14,674 ± 1247 pM, n = 3) in the CMC spot). All in all this demonstrated that the tools for determination of low abundance biomarkers at upper reference level from dried matrix spots now is available through a combination of immunoaffinity enrichment and state-of-the-art LC-MS/MS.

Dispersion-assisted quick and simultaneous extraction of 30 pesticides from alcoholic and non-alcoholic drinks with the aid of experimental design.

The presence of pesticides in food items and beverages is a big threat to humankind, and their quantitative estimation with high precision and accuracy is always a challenge for analytical chemists. Hence, a simple and rapid method is proposed for the simultaneous determination of 30 pesticides in beverages (alcoholic and non-alcoholic drinks). The proposed method hyphenated with triple quadrupole liquid chromatography mass spectrometry has only 2min chromatographic runtime for the analysis of all the pesticides. All the factors affecting the extraction yield have been optimized using an experimental design; and under optimized conditions, the developed method has been validated. The detection limits for all the pesticides were in the range of 0.001-0.348μg/L with good linearity in the concentration range of 0.01-80.0μg/L. The coefficient of determination was in the range of (R2 )≥0.977 to 0.999 for all the pesticides. The method was also checked for the precision of the relative standard deviation, which was below 4.75 (intra-day) and 8.96% (inter-day). The recovery of the method was 92-138%.

Glyphosate residues in Swiss market foods: monitoring and risk evaluation

ABSTRACTA total of 243 samples of diverse foodstuffs were analysed for glyphosate and aminomethylphosphonic acid (AMPA) using a liquid chromatography triple quadrupole mass spectrometry (LC/MS/MS) method with a relatively low limit of quantification in the range of 0.0005–0.0025 mg kg−1. Main contributors for dietary glyphosate and AMPA intake were cereals and pulses. The results suggest that pasta is a very important foodstuff for dietary glyphosate residue intake in Switzerland. Interestingly all samples of wine, fruit juice and nearly all samples of honey tested positive for glyphosate although at very low levels. A dietary risk assessment was conducted. Food products for analysis were not selected purely at random, rather products were selected for which high levels of glyphosate residues were suspected. However, even in samples where high residue levels were expected, no exceedances of maximum residue levels were found. Consequently, human exposure did not exceed neither acceptable daily intake nor acute reference dose. Therefore, glyphosate residues found in the sampled foodstuffs from the Swiss market were of no concern for human health.

Development of a simple and sensitive liquid chromatography triple quadrupole mass spectrometry (LC–MS/MS) method for the determination of cannabidiol (CBD), Δ9-tetrahydrocannabinol (THC) and its metabolites in rat whole blood after oral administration of a single high dose of CBD

The investigation of the possible conversion of cannabidiol (CBD) into Δ9-tetrahydrocannabinol (THC) in vivo after oral administration of CBD is reported herein since recent publications suggested a rapid conversion in simulated gastric fluid. To this end, single high dose of CBD (50mg/kg) was administered orally to rats and their blood was collected after 3 and 6h. A highly sensitive and selective LC–MS/MS method was developed and fully validated in compliance with the Scientific Working Group of Forensic Toxicology (SWGTOX) standard practices for method validation in forensic toxicology. This method also involved the optimization of cannabinoids and their metabolites extraction in order to remove co-eluting phospholipids and increase the sensitivity of the MS detection. Neither THC nor its metabolites were detected in rat whole blood after 3 or 6h from CBD administration. After oral administration, the amount of CBD dissolved in olive oil was higher than that absorbed from an ethanolic solution. This could be explained by the protection of lipid excipients towards CBD from acidic gastric juice.

Effects of sulfur-fumigation on the pharmacokinetics, metabolites and analgesic activity of Radix Paeoniae Alba

Ethnopharmacological relevanceRadix Paeoniae Alba (Baishao, BS), one of the most commonly used traditional Chinese medicinal herbs, has many pharmacological effects including analgesic activity. Previous studies found that sulfur-fumigation, a post-harvest handling process developed to prevent mold contamination of medicinal herbs, altered the quality of BS. However, whether sulfur-fumigation affects the pharmacokinetics, safety and efficacy of BS warrants further investigation. Aim of the studyTo evaluate the feasibility of sulfur-fumigation as a post-harvest handling process of BS from the viewpoints of pharmacokinetics, safety and efficacy. Materials and methodsThe pharmacokinetic behaviors of four active components of BS and one characteristic component of sulfur-fumigated BS (S-BS) were evaluated by high performance liquid chromatography triple quadrupole mass spectrometry (HPLC-TQ-MS/MS). The safety was investigated using ultra high performance liquid chromatography quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-MS/MS) based metabolomics approach after intragastric (i.g.) administration of non-fumigated BS (N-BS) and S-BS in rats. The analgesic efficacy was compared using hot-plate test in mice, after i.g. administration of N-BS and S-BS, at both high and low dosages. ResultsSystemic exposures of paeoniflorin and oxypaeoniflorin, two analgesic components of BS, were significantly decreased in the S-BS treated group compared to the N-BS treated group, while paeoniflorin sulfonate, one of the sulfur-containing derivatives of S-BS, was detected in all time-points of S-BS treated group with the area under the plasma concentration–time curve (AUC0−t) and the maximum plasma concentration (Cmax) as high as 7077.06 ± 2232.97ng/mL*h and 1641.42 ± 634.79ng/mL respectively, which indicated that sulfur-fumigation altered the pharmacokinetic behaviors of BS. Besides, paeoniflorin sulfonate and its four metabolites with ambiguous toxicities, as well as one endogenous metabolite p-cresol glucuronide, the biomarker of disordered homeostasis of intestinal bacteria and bile acid, were identified as the characteristic metabolites in S-BS administered rats, suggesting that sulfur-fumigation reduced the safety of BS. Furthermore, the analgesic effects at both low and high dosages were decreased in different extent when compared to N-BS administered groups, indicating that sulfur-fumigation weakened the efficacy of BS. ConclusionSulfur-fumigation altered the pharmacokinetics, as well as reduced the safety and efficacy of BS, suggesting that sulfur-fumigation is not recommended for post-harvest handling of BS.

Bioactive small molecules in commercially available cereal food: Benzoxazinoids

The presence of bioactive benzoxazinoids (BXs) in mature whole grain and experimental bread products was recently shown. The aim of this study was to reveal the level of BXs in a broad range of commercial cereal products and discuss these findings in relation to ingredients and preparation processes. The concentrations of 11 BX compounds were determined by liquid chromatography–triple quadrupole mass spectrometry (LC–MS/MS) in 25 supermarket-available cereal food products. The total BX content varied from below the limit of detection to >500μg/g dry matter (DM) with some degree of variation among different lots. The products W (Rugsandwich), H (Gulerodsrugbrød), L (Mørke Multikerneboller) and AA (Solsikkeklapper), which were all rich in rye-based ingredients, had a total BX concentration >300μg/g DM. Double hexose derivatives of 2-hydroxy-1,4-benzoxazin-3-one (HBOA), 2,4-dihydroxy-1,4-benzoxazin-3-one (DIBOA) and 2-β-d-glucopyranosyloxy-4-hydroxy-1,4-benzoxazin-3-one (DIBOA-glc) were the major BXs in most products. Further studies into the health effects of BXs should be conducted, and future exploitation of these compounds seems possible.

Simultaneous determination of acrylamide and 4-hydroxy-2,5-dimethyl-3(2H)-furanone in baby food by liquid chromatography–tandem mass spectrometry

A liquid chromatography triple quadrupole mass spectrometry method was developed and validated for the simultaneous determination of acrylamide and 4-hydroxy-2,5-dimethyl-3(2H)-furanone (HDMF) in baby food. The sample preparation involves acetonitrile-based extraction combined with dispersive primary secondary amine (PSA) cleanup and cation-exchange solid-phase extraction (SPE), which promotes efficient removal of matrix interferences. Analytical selectivity and sensitivity were achieved for the quantification of acrylamide and HDMF in complex matrices such as fruit, cereal and milk-based baby foods; furthermore, adequate linearity (range 10–300μgkg−1) in solvent and matrix-matched calibration curves, and appropriate recoveries (94–110%) and precision (RSD≤10%), under repeatability and within-laboratory reproducibility conditions, were also obtained. Expanded measurement uncertainty was estimated at the 20μgkg−1 level (limit of quantification) on the basis of data obtained from in-house validation, with values of 25.5 and 16.5% for acrylamide and HDMF, respectively. The fitness for purpose of developed method was verified by analyzing 15 commercial baby foods available in the Brazilian market. Acrylamide was detected in one plum-based baby food (35μgkg−1) while HDMF in 67% of the samples analyzed (levels between 25 and 262μgkg−1).

Germline BAP1 mutations induce a Warburg effect

Carriers of heterozygous germline BAP1 mutations (BAP1+/−) develop cancer. We studied plasma from 16 BAP1+/− individuals from 2 families carrying different germline BAP1 mutations and 30 BAP1 wild-type (BAP1WT) controls from these same families. Plasma samples were analyzed by liquid chromatography time-of-flight mass spectrometry (LC-TOF-MS), ultra-performance liquid chromatography triple quadrupole mass spectrometry (UPLC-TQ-MS), and gas chromatography time-of-flight mass spectrometry (GC-TOF-MS). We found a clear separation in the metabolic profile between BAP1WT and BAP1+/− individuals. We confirmed the specificity of the data in vitro using 12 cell cultures of primary fibroblasts we derived from skin punch biopsies from 12/46 of these same individuals, 6 BAP1+/− carriers and 6 controls from both families. BAP1+/− fibroblasts displayed increased aerobic glycolysis and lactate secretion, and reduced mitochondrial respiration and ATP production compared with BAP1WT. siRNA-mediated downregulation of BAP1 in primary BAP1WT fibroblasts and in primary human mesothelial cells, led to the same reduced mitochondrial respiration and increased aerobic glycolysis as we detected in primary fibroblasts from carriers of BAP1+/− mutations. The plasma and cell culture results were highly reproducible and were specifically and only linked to BAP1 status and not to gender, age or family, or cell type, and required an intact BAP1 catalytic activity. Accordingly, we were able to build a metabolomic model capable of predicting BAP1 status with 100% accuracy using data from human plasma. Our data provide the first experimental evidence supporting the hypothesis that aerobic glycolysis, also known as the ‘Warburg effect’, does not necessarily occur as an adaptive process that is consequence of carcinogenesis, but rather that it may also predate malignancy by many years and facilitate carcinogenesis.

Simultaneous determination of pyrethroids and pyrethrins by dispersive liquid-liquid microextraction and liquid chromatography triple quadrupole mass spectrometry in environmental samples.

A simple and environmentally friendly dispersive liquid-liquid microextraction (DLLME) method coupled with electrospray ionization liquid chromatography triple quadrupole mass spectrometry (LC-QqQ-MS/MS) was developed for the simultaneous determination of 17 synthetic and natural pyrethroids. A comparison of solid-phase extraction (SPE) versus DLLME for water samples and only "dilute and shoot" versus the additional extract cleanup by DLLME for sediment samples is reported. Chloroform was the extracting solvent in the DLLME technique for both water and sediment samples. Ultrasonic energy was applied to fully extract the analytes into fine droplets, providing high recoveries in short times. Method detection limits (MDLs) ranged from 0.12 to 0.62ngL-1 and recoveries from 70 to 119% with RSD values 2-15% (n=5) for water samples. In sediment samples, MDLs ranged from 0.50 to 2.50ngg-1 and recoveries from 71 to 112% with RSDs 2-16% (n=5). The proposed method showed a good linearity within the range of 10-500ngmL-1, with coefficients of determination (R 2) higher than 0.99. Matrix effects were observed for most compounds in water and sediment (ME%<-10%). The proposed methodology was applied for the analysis of water and sediment samples from Albufera wetland and Turia River. Acrinathrin (48ngg-1) and etofenprox (16ngg-1) were detected in sediment samples. Graphical abstract Pyrethrins and pyrethroid determination in water and sediments by DLLME and LC-MS/MS.

Analysis of toldimfos in porcine muscle and bovine milk using liquid chromatography-triple quadrupole mass spectrometry.

An analytical method was developed for the detection of toldimfos sodium residues in porcine muscle and bovine milk using liquid chromatography-triple quadrupole tandem mass spectrometry (LC-MS/MS) analysis. The drug was extracted from muscle and milk using 10 mm ammonium formate in acetonitrile and then purified using n-hexane. The drug was well separated on a Luna C18 column using a mixture of 10 mm ammonium formate in ultrapure water (A) and acetonitrile (B) as the mobile phase. Good linearity was achieved over the tested concentration range (0.005-0.03 mg/kg) in matrix-matched standard calibration. The determination coefficients (R2 ) were 0.9942 and 0.9898 for muscle and milk, respectively. Fortified porcine muscle and bovine milk contained concentrations equivalent to and twice the limit of quantification (0.005 mg/kg) yielded recoveries in the range of 75.58-89.74% and relative standard deviations of ≤8.87%. Samples collected from large markets located in Seoul, Republic of Korea, tested negative for toldimfos sodium residue. In conclusion, ammonium formate in acetonitrile can effectively extract toldimfos sodium from porcine muscle and bovine milk without solid-phase extraction, which is usually required for cleanup before analysis. This method can be applied for the routine analysis of toldimfos in foods of animal origins.

Comparison of green sample preparation techniques in the analysis of pyrethrins and pyrethroids in baby food by liquid chromatography–tandem mass spectrometry

A new selective and sensitive liquid chromatography triple quadrupole mass spectrometry method was developed for simultaneous analysis of natural pyrethrins and synthetic pyrethroids residues in baby food. In this study, two sample preparation methods based on ultrasound-assisted dispersive liquid–liquid microextraction (UA-DLLME) and salting-out assisted liquid–liquid extraction (SALLE) were optimized, and then, compared regarding the performance criteria. Appropriate linearity in solvent and matrix-based calibrations, and suitable recoveries (75–120%) and precision (RSD values≤16%) were achieved for selected analytes by any of the sample preparation procedures. Both methods provided the analytical selectivity required for the monitoring of the insecticides in fruit-, cereal- and milk-based baby foods. SALLE, recognized by cost-effectiveness, and simple and fast execution, provided a lower enrichment factor, consequently, higher limits of quantification (LOQs) were obtained. Some of them too high to meet the strict legislation regarding baby food. Nonetheless, the combination of ultrasound and DLLME also resulted in a high sample throughput and environmental-friendly method, whose LOQs were lower than the default maximum residue limit (MRL) of 10μgkg−1 set by European Community for baby foods. In the commercial baby foods analyzed, cyhalothrin and etofenprox were detected in different samples, demonstrating the suitability of proposed method for baby food control.

Multiresidue determination of estrogens in different dairy products by ultra-high-performance liquid chromatography triple quadrupole mass spectrometry

In this work, a simple and fast methodology has been validated and applied for the analysis of a group of 22 estrogenic compounds including eight phytoestrogens (i.e. daidzein, enterodiol, glycitein, enterolactone, genistein, formononetin, prunetin, biochanin A), six mycotoxins (β-zearalanol, β-zearalenol, α-zearalanol, α-zearalenol, zearalanone, zearalenone) as well as four synthetic (i.e. ethynylestradiol, diethylstilbestrol, dienestrol, hexestrol) and four natural estrogens (i.e. estriol, 17β-estradiol, 17α-estradiol, estrone) in different dairy products. Extraction was carried out using the QuEChERS method while separation, determination and quantification of the target analytes were achieved by ultra-high-performance liquid chromatography coupled to triple quadrupole mass spectrometry with an electrospray ionization interface. The methodology was validated for four dairy product samples with relevant interest for the population including skimmed and whole cheese and goat and cow kefir, using 17β-estradiol-2,4,16,16,17-d5 as internal standard for natural and synthetic estrogens and β-zeralanol-10,10,11,12,12-d5 as internal standard for mycotoxins and phytoestrogens. Recovery ranged from 70 to 119% for the four types of matrices with RSD values lower than 14% and the limits of quantification of the method achieved were in the range 0.025–2.50μg/kg for all samples. Finally, the analysis of commercially available products was carried out finding the presence of daidzein, glycitein enterolactone and genistein in some of the studied samples.

Serum lipid alterations identified in chronic hepatitis B, hepatitis B virus-associated cirrhosis and carcinoma patients

The incidences of chronic hepatitis B (CHB), Hepatitis B virus (HBV)-associated cirrhosis and HBV-associated carcinoma are high and increasing. This study was designed to evaluate serum lipid metabolite changes that are associated with the progression from CHB to HBV-associated cirrhosis and ultimately to HBV-associated HCC. A targeted metabolomic assay was performed in fasting sera from 136 CHB patients, 104 HBV-associated cirrhosis, and 95 HBV-associated HCC using ultra-performance liquid chromatography triple quadrupole mass spectrometry. A total of 140 metabolites were identified. Clear separations between each two groups were obtained using the partial least squares discriminate analysis of 9 lipid metabolites. Progressively lower levels of long-chain lysophosphatidylcholines (lysoPC a C18:2, lysoPC a C20:3, lysoPC a C20:4) were observed from CHB to cirrhosis to carcinoma; lower levels of lysoPC a C20:4 were found in patients with higher model for end-stage liver disease in the same disease group; and lysoPC a C20:3 levels were lower in Child-Pugh Class C than in Class A and Class B in HBV-associated cirrhosis and HBV-associated HCC groups. The octadecadienyl carnitine level was higher in HBV-associated cirrhosis group than in other two groups. Serum levels of selected long-chain lysoPCs are promising markers for the progression of HBV-associated liver diseases.