Low-Temperature Cleanup Followed by Dispersive Solid-Phase Extraction for Determination of Nine Polar Plant Growth Regulators in Herbal Matrices Using Liquid Chromatography-Tandem Mass Spectrometry.

Polar plant growth regulators, used alone or doped in fertilizers, are most effective and widely utilized plant growth regulators (PGRs) in agriculture, which play important roles in mediating the yield and quality of crops and foodstuffs. The application scope has been extended to herbal medicines in the past 2 decades and relevant study is inadequate. The aim of this study is to establish a QuPPe-based extraction method containing low-temperature and d-SPE cleanup procedure followed by the detection on a selective multiresidue ultrahigh-performance liquid chromatography - triple quadrupole tandem mass spectrometry (UHPLC-QqQ-MS/MS) in three herbal matrices. This simple, accurate, versatile and robust method was verified according to the validation criteria of the SANTE/12682/2019 guideline document. The analytical range was from 2.5 to 200 µg/L, and the average recoveries were in the range of 64.6-117.8% (n = 6). The optimized method was applied to 135 herbal medicines thereof. Result showed that the detection frequency of chlormequat was the highest in the investigated PGRs, with the positive rate of 15.6%. Improvement of the detection method for polar PGRs will enrich the coverage of PGRs, which is conducive to safeguard public health and ensure drug safety. Supplementary Information: The online version contains supplementary material available at 10.1007/s10337-023-04254-3.

[Component profile analysis of Sanhan Huashi Formula based on UHPLC-LTQ-Orbitrap-MS, GC-MS, and UHPLC-QQQ-MS/MS].

This study comprehensively analyzed the active components of Sanhan Huashi Formula using qualitative and quantitative mass spectrometry techniques, laying the foundation for understanding its pharmacological substance basis. UHPLC-LTQ-Orbitrap-MS and GC-MS technologies were used to analyze and identify the volatile and non-volatile components in Sanhan Huashi Formula. UHPLC-QQQ-MS/MS technology was used to simultaneously determine the content of 27 major active components in the formula. The results showed that 308 major chemical components were identified in Sanhan Huashi Formula, among which 60 compounds were identified by comparing with reference standards, mainly including alkaloids, flavonoids, coumarins, triterpenoid saponins, amino acids, and nucleosides. GC-MS technology preliminarily identified 52 volatile compounds, with γ-eudesmol and ß-eudesmol as the main components. The quantitative results demonstrated good linearity(r>0.99) for the 27 active components, indicating the stability, simplicity, and reliability of the established method. Among them, amygdalin, nodakenin, arecoline, ephedrine, and pseudoephedrine had relatively high content and were presumably the main pharmacologically active substances. In conclusion, this study systematically and comprehensively characterized the major chemical components and patterns in Sanhan Huashi Formula, providing a basis for understanding its pharmacological mechanisms and clinical applications.

Metabolite profiling analysis of plasma, urine, and feces of rats after oral administration of Flos Chrysanthemi Indici preparation through UHPLC-Q-Exactive-MS combined with pharmacokinetic study of markers by UHPLC-QQQ-MS/MS.

Zhenju Antihypertensive Tablet is one of the Flos Chrysanthemi Indici preparations (FCIP) with antihypertensive effect. In order to investigate its process in vivo, a method using ultra-high performance liquid chromatography coupled with quadruple Exactive mass spectrometry (UHPLC-Q-Exactive-MS) was established for comprehensive analysis of the absorbed active component and metabolites in rat plasma, urine, and feces after the oral administration of FCIP. As a result, a total of 61 FCIP-related xenobiotics were identified, including 35 in plasma (25 prototypes, 10 metabolites), 40 in urine (23 prototypes, 17 metabolites), and 25 in feces (12 prototypes, 13 metabolites). The metabolism reactions included phase I reactions (such as oxidation, methylation, and reduction) and phase II reactions (such as sulfate conjugation and glucuronide conjugation). Meanwhile, a verified and optimized ultra-performance liquid chromatography with triple quadrupole mass spectrometry (UHPLC-QQQ-MS/MS) method was developed for the simultaneous investigation of the pharmacokinetic profiles of four markers in FCIP. The results showed that all of the four components had good oral absorption effect. This study simultaneously investigated the comprehensive metabolic profiling of FCIP in rat plasma, urine, and feces after the oral administration as well as the four components pharmacokinetic behavior. The results can provide the therapeutic material basis for FCIP.

Determination of Mifepristone (RU-486) and Its Metabolites in Maternal Blood Sample after Pharmacological Abortion.

The aim of the study was the development and validation of the UHPLC-QqQ-MS/MS method for the determination of mifepristone in human blood as well as the identification and quantification of its metabolites after self-induced pharmacological abortion. The metabolic pathway in humans was proposed after examination of an authentic casework. The fast and simple preanalytical procedure was successfully applied (pH9, tert-butyl-methyl ether). The validation parameters of the method were as follows: limit of quantification: 0.5 ng/mL; coefficients of determination: >0.999 (R2), intra- and inter-day accuracy and precision values did not exceed ± 13.2%. The recovery and matrix effect were in the range of 96.3−114.7% and from −3.0 to 14.7%, respectively. Toxicological analysis of the mother's blood (collected the day after the pregnancy termination) revealed the presence of five compounds: mifepristone (557.4 ng/mL), N-desmethyl-mifepristone (638.7 ng/mL), 22-OH-mifepristone (176.9 ng/mL), N,N-didesmethyl-mifepristone (144.5 ng/mL) and N-desmethyl-hydroxy-mifepristone (qualitatively). To our knowledge, the study presented in this paper is the first report on the concentrations of mifepristone and its metabolites in maternal blood samples after performing a self-induced abortion. The established UHPLC-QqQ-MS/MS method is suitable for forensic toxicological analysis as well as in terms of clinical toxicology in future investigations (examination of pharmacokinetics, bioavailability and metabolism of RU-486).

Forensic Toxicological Aspects of Misoprostol Use in Pharmacological Abortions.

The aim of this study was establishment of an UHPLC-QqQ-MS/MS method for the deter-mination of misoprostol acid in biological specimens in cases of pharmacological abortions. Forensic toxicological examination was performed in three different biological samples (whole blood, placenta and fetal liver). The validation parameters of the method were as follows: limit of detection: 25 pg/mL; limit of quantification: 50 pg/mL, coefficient of determination: >0.999 (R2), intra- and interday accuracy and precision: not greater than 13.7%. The recovery and matrix effect were in the range of 88.3−95.1% and from −11.7 to −4.9%, respectively. Toxicological analysis of the mother's blood (collected two days after pregnancy termination) did not reveal any abortifacients; however, misoprostol acid was found in the placenta (793 pg/g) and fetal liver (309 pg/g). The second case involved a fetus found near a garbage container. The concentration of misoprostol acid in the placenta was 2332 pg/g. In the presented study, an extensive literature review of misoprostol pharmacokinetics studies was performed. To our knowledge, the UHPLC-QqQ-MS/MS technique presented in this paper is the first quantitative method applied for forensic toxicological purposes. In addition, postmortem concentrations of misoprostol acid in miscarried fetuses due to illegal abortions were reported for the first time.

Phytoprostanes, phytofurans, tocopherols, tocotrienols, carotenoids and free amino acids and biological potential of sea buckthorn juices.

BACKGROUND: Juices are currently a fast-growing segment in the fruit and vegetable industry sector. However, there are still no reports on the diversity of the phytochemical profile and health-promoting properties of commercial sea buckthorn (Hippophaë rhamnoides) juices. This study aimed to identify and quantify phytoprostanes, phytofurans by ultra high-performance liquid chromatography coupled with triple quadrupole mass spectrometry (UHPLC-QqQ-MS/MS), tocopherols, tocotrienols by ultra-performance liquid chromatography coupled with a fluorescence detector (UPLC-FL), carotenoids, and free amino acids by ultra-performance liquid chromatography coupled with a photodiode detector-quadrupole and tandem time-of-flight mass spectrometry (UPLC-PDA-Q/TOF-MS), and assess their anti-cholinergic, anti-diabetic, anti-obesity, anti-inflammatory, and antioxidant potential by in vitro assays of commercial sea buckthorn juices. RESULTS: Phytoprostanes (PhytoPs) and phytofurans (PhytoFs) in sea buckthorn juices were identified for the first time. Juices contained eight F1 -, D1 -, B1 - and L1 -phytoprostanes and one phytofuran (32.31-1523.51 ng and up to 101.47 µg/100 g dry weight (DW)), four tocopherol congeners (22.23-94.08 mg 100 g-1 DW) and three tocotrienols (5.93-25.34 mg 100 g-1 DW). Eighteen carotenoids were identified, including ten xanthophylls, seven carotenes and phytofluene, at a concentration of 133.65 to 839.89 mg 100 g-1 DW. Among the 20 amino acids (175.92-1822.60 mg 100 g-1 DW), asparagine was dominant, and essential and conditionally essential amino acids constituted 11 to 41% of the total. The anti-enzyme and antioxidant potential of juices correlated selectively with the composition. CONCLUSION: Sea buckthorn juice can be a valuable dietary source of vitamins E and A, oxylipins and amino acids, used in the prevention of metabolic syndrome, inflammation, and neurodegenerative processes. The differentiation of the composition and the bioactive potential of commercial juices indicate that, for the consumer, it should be important to choose juices from the declared berry cultivars and crops. © 2021 Society of Chemical Industry.

Discovery of quality control ingredients in burdock root by combining anti-tumor effects and UHPLC-QqQ-MS/MS.

Burdock root is the root of Arctium lappa L., a plant of the Compositae family, which has the effects of dispersing wind and heat, detoxifying and reducing swelling. In order to better control the quality of burdock root, a screening study of quality control indicators was carried out. The current research combines biological activity evaluation with chemical analysis to screen and identify the biologically active compounds of burdock root as chemical components for the quality control of herbal medicine. The efficacy of 10 batches of ethanol extracts of burdock roots was evaluated by a tumor inhibition experiment in S180 tumor-bearing mice. The five main chemical components of these extracts were simultaneously quantitatively measured by ultra-high performance liquid chromatography combined with triple quadrupole mass spectrometry. Pearson correlation analysis was used to establish the relationship between these extracts' biological activity and chemical properties. The results showed that chlorogenic acid, caffeic acid and cynarin were positively correlated with the effect of inhibiting tumor growth, and further bioassays confirmed this conclusion. In conclusion, chlorogenic acid, caffeic acid and cynarin can be used as quality control markers for burdock root's antitumor effect.

Determination of pesticide residues in dates using UHPLC-QqQ-MS/MS: method development and validation.

A modified, efficient, and sensitive acetate-buffered QuEChERS extraction method was developed for the quantitative study of 16 commonly applied multiclass pesticides on date palm fruit. The date palm fruit samples were rehydrated by adding water during comminution. Samples were extracted with acidified acetonitrile, buffered with acetate salt. To minimize the matrix interferences, clean-up of the rehydrated samples was optimized by comparison with different sorbents (alumina, silica gel, florisil, primary secondary amine (PSA), and chitosan). The method validation parameters were evaluated as per European Union (EU) guidelines (SANTE/12682/2019). For 16 pesticides, % recovery of 69 to 121.8% with an associated precision (RSD ≤ 20%) was achieved at the fortification levels that were 0.5 to 2 times of European Union maximum residue limits (EU-MRLs). The validated method was successfully employed for the analysis of date palm fruit samples (n = 20) collected from various markets. Forty percent (40%) of samples (n = 8) were found to be contaminated with various pesticides. The most frequently detected residues were carbofuran, carbaryl, metalaxyl, tebuconazole, triazophos, and pyriproxyfen. The concentration of all the detected pesticides in real samples was below the EU-MRLs.

Quantification of Nineteen Bioactive Components in the Ancient Classical Chinese Medicine Formula of Wen-Dan Decoction and Its Commercial Preparations by UHPLC-QQQ-MS/MS.

A UHPLC-QQQ-MS/MS method was developed to quantify the significant constituents in Wen-Dan Decoction (WDD), a traditional Chinese medicine. Analysis of 19 compounds was conducted on an ACQUITY UPLC® BEH C18 Column (2.1 × 50 mm, 1.7 µm) using elution with a gradient elution of acetonitrile and 0.05% (v/v) formic acid in water. A triple quadrupole mass spectrometer was operated in negative ionization mode and positive ionization mode by multiple reaction monitoring (MRM), respectively. All calibration curves showed acceptable linearity (r ≥ 0.9950). The RSDs of intra- and inter-day precisions of low, mid and high concentrations were ≤ 8.88%. The repeatabilities (RSDs ≤ 7.17%) and stabilities (RSD ≤ 4.79%) of the samples were qualified. The recoveries were found in the range of 93.07 ± 3.86 to 103.98 ± 2.98% with the RSD varying between 1.30 and 7.86%. The final rapid, sensitive, precise, accurate and reliable UHPLC-QQQ-MS/MS method was used for the simultaneous quantification of 19 constituents in WDD and its commercial preparations. The strategy of combining the contents of the 19 chemicals in a daily dose of the WDD preparations with the hierarchical cluster analysis and the 3D principal component analysis was employed to effectively distinguish the WDD preparations provided by the different suppliers, which represents a contribution to the evaluation and control of the quality of WDD (or other decoctions consisting of the same herbs) and the preparations of WDD in other dosage forms such as tablets and granules.

Correlation between Quality and Geographical Origins of Poria cocos Revealed by Qualitative Fingerprint Profiling and Quantitative Determination of Triterpenoid Acids.

Poria cocos (Schw.) Wolf (PC) is a well-known saprophytic fungus, and its sclerotium without the epidermis (PCS) is widely used in traditional Chinese medicine and as a functional food in many countries. PCS is normally collected from multiple geographical regions, but whether and how the quality of PCS correlates with where it grows have not been determined. This correlation could be significant both for quality control and optimum utilization of PCS as a natural resource. In this study, a qualitative fingerprint profiling method performed by ultra-performance liquid chromatography (UHPLC) with diode array detection (DAD) combining quadrupole time-of-flight-mass spectrometry (QTOF-MS/MS) and a quantitative UHPLC coupled with triple quadrupole mass spectrometry (QqQ-MS/MS) approach were established to investigate whether and how the quality of PCS correlates with its collection location. A standard fingerprint of PCS was generated by median simulation of 25 tested samples collected from four main producing areas of China, and similarity analysis was applied to evaluate the similarities between the fingerprints of samples and the standard fingerprint. Twenty three common peaks occurring in the fingerprint were unequivocally or tentatively identified by UHPLC-QTOF-MS/MS. Meanwhile, principal component analysis (PCA), supervised orthogonal partial least squares-discriminate analysis (OPLS-DA) and hierarchical cluster analysis (HCA) were employed to classify 25 batches of PCS samples into four groups, which were highly consistent with the four geographical regions. Ten compounds were screened out as potential markers to distinguish the quality of PCS. Nine triterpene acids, including five compounds that played important roles in the clusters between different samples collected from the four collection locations, were simultaneously quantified by using the multiple reaction monitoring (MRM) mode of UHPLC-QqQ-MS/MS. The current strategy not only clearly expounded the correlation between quality and geographical origins of PCS, but also provided a fast, accurate and comprehensive qualitative and quantitative method for assessing the quality of PCS.

Determination of several families of phytochemicals in different pre-cooked convenience vegetables: effect of lifetime and cooking.

Phytochemicals content, including several families such as phenolic acids, isoflavones, flavones, flavonols, isothiocyanates, and glucosinolates, was determined in pre-cooked convenience vegetables by ultra high performance liquid chromatography coupled to triple quadrupole tandem mass spectrometry (UHPLC-QqQ-MS/MS). It was observed that there is not a common behavior of the individual concentration of phytochemicals during the lifetime and cooking of the matrix, and compounds change their concentration without a specific trend. It was observed that neither lifetime nor cooking process have significant effects on the total content of phytochemicals except in broccoli, although some changes in the individual content of the target compounds were observed, suggesting that interconversion processes could be performed during the lifetime and/or cooking process of the product.

Development of a targeted method for DNA adductome and its application as sensitive biomarkers of ambient air pollution exposure.

DNA adducts are widely recognized as biomarkers of exposure to environmental carcinogens and associated health effects in toxicological and epidemiological studies. This study presents a targeted and sensitive method for comprehensive DNA adductome analysis using ultra-high-performance liquid chromatography coupled with triple-quadrupole tandem mass spectrometry (UHPLC-QqQ-MS/MS). The method was developed using calf thymus DNA, with careful optimization of mass spectrometric parameters, chromatographic separation conditions, and pretreatment methods. Ultimately, a targeted method was established for 41 DNA adducts, which showed good linearity (R2 ≥0.992), recovery (80.1-119.4 %), accuracy (81.3-117.8 %), and precision (relative standard deviation <14.2 %). The established method was employed to analyze DNA adducts in peripheral blood cells from pregnant women in Shanxi and Beijing. Up to 23 DNA adducts were successfully detected in samples of varying sizes. From 2 µg of maternal DNA samples, seven specific adducts were identified: 5-methyl-2'-deoxycytidine (5-MedC), 5-hydroxymethyl-2'-deoxycytidine (5-HmdC), N6-methyl-2'-deoxyadenosine (N6-MedA), 8-hydroxy-2'-deoxyguanosine (8-OHdG), 5-hydroxy-2'-deoxycytidine (5-OHdC), 1,N6-etheno-2'-deoxyadenosine (1,N6-εdA), and N2-methyl-2'-deoxyguanosine (N2-MedG). This study reveals that exposure to higher concentrations of ambient air pollutants may elevate the levels of DNA methylation and oxidative damage at different base sites, highlighting the application potential of DNA adducts as sensitive biomarkers of air pollution exposure.

"Plant Golden" C. sativus: Qualitative and quantitative analysis of major components in stigmas and petals and their biological activity in vitro.

Crocus sativus L. (C. sativus) has its stigma as the main valuable part used. With extremely low production and high prices, stigma is considered a scarce resource. As a result, its petals, considered as by-products, are often discarded, leading to significant waste. We developed a UPLC-Q-Orbitrap HRMS method for qualitative analysis of stigmas and petals and a UHPLC-QQQ-MS/MS method for simultaneous quantification of 9 characteristic active compounds for the first time, and compared their biological activity in vitro. The results indicated that a total of 63 compounds were identified in the petals and stigmas. The content of flavonoids in the petals was significantly superior to that in the stigma, and the content of quercetin in the petals was 50 times higher than that in the stigma. The results of the in vitro evaluation of biological activity indicated that both the petals （•OH: IC50=39.70 mg/mL; DPPH: IC50=28.37 mg/mL; ABTS: IC50=0.9868 mg/mL）and stigma （•OH: IC50=34.41 mg/mL; DPPH: IC50=38.99 mg/mL; ABTS: IC50=3.194 mg/mL）demonstrated comparable antioxidant activities. However, the tyrosinase inhibitory activity in petals （IC50=21.17 mg/mL） was weaker than that in stigma（IC50=1.488 mg/mL）. This study provides a fast, reliable, and efficient analytical method that can be used for the quality assessment of petals as a natural resource and its related products in the food and pharmaceutical industries.

Cross-sectional, commercial testing, and chromatographic study of the occurrence of antibiotic residues throughout an artisanal raw milk cheese production chain.

This study investigated antibiotic utilization in artisanal dairies and residue occurrence throughout the raw milk cheese production chain using commercial testing (Charm KIS and Eclipse Farm3G) and UHPLC-QqQ-MS/MS and LC-QqQ-MS/MS. The cross-sectional survey results revealed gaps in the producers' knowledge of antibiotic use. Commercial testing detected antibiotic levels close to the LOD in 12.5 % of the samples, mainly in raw milk and whey, with 10.0 % testing positive, specifically in fresh and ripened cheeses, indicating that antibiotics are concentrated during cheese-making. Chromatographically, several antibiotics were identified in the faeces of healthy animals, with chlortetracycline (15.7 ± 34.5 µg/kg) and sulfamethazine (7.69 ± 16.5 µg/kg) predominating. However, only tylosin was identified in raw milk (3.28 ± 7.44 µg/kg) and whey (2.91 ± 6.55 µg/kg), and none were found in fresh or ripened cheeses. The discrepancy between commercial and analytical approaches is attributed to compounds or metabolites not covered chromatographically.

Phenolic Composition of Red and White Wine Byproducts from Different Grapevine Cultivars from La Rioja (Spain) and How This Is Affected by the Winemaking Process.

The recovery of raw materials offers an opportunity for applying the principles of circular bioeconomy. The phenolic composition of three underused wine byproducts (skin, seed, and bunch stem) was analyzed through UHPLC-QqQ-MS/MS to evaluate the intercultivar variability comparing red and white grape cultivars from La Rioja (Spain) and the influence of the winemaking, comparing conventional fermentation and carbonic maceration. We observed that the red skin, especially from Graciano, is rich in anthocyanins, whereas the white skin contains mainly phenolic acids, flavonols, and flavan-3-ols, with Maturana Blanca being the richest variety. Seeds are rich in flavan-3-ols and lignans with Maturana Blanca and Viura, respectively, the richest cultivars. Stems contain high amounts of flavan-3-ols, lignans, and stilbenes, with the red cultivars of Garnacha and Tempranillo being the richest samples. Carbonic maceration has a negative effect on the phenolic amount compared to conventional fermentation. In synthesis, we observed that each type of byproduct from red or white grape cultivars has a particular phenolic composition that can result in obtaining different ingredients with particular phenolic composition for target applications.

Determination of Prostaglandins (Carboprost, Cloprostenol, Dinoprost, Dinoprostone, Misoprostol, Sulprostone) by UHPLC-MS/MS in Toxicological Investigations.

Prostaglandins have stimulative influence on the human uterus and therefore were introduced to medical treatment in reproductive healthcare as labor inductors or abortifacients. The UHPLC-ESI-QqQ-MS/MS method was developed for six prostaglandins: carboprost, cloprostenol, dinoprost (PGF2α), dinoprostone (PGE2), misoprostol and sulprostone (substances for pregnancy termination) in pharmaceutical samples and was applied for the toxicological examination of pills containing misoprostol (collected during gynecological examination). There were used two internal standards: misoprostol-d5 and PGF2α-d4. The quantification of analytes was performed in the MRM mode. The linearity of method was in the range from 0.1 to 10 µg/mL, with a coefficient of determination above 0.997 (R2) for each compound. The precision and accuracy values did not exceed ±5.0%. Analysis of the pills revealed the presence of two substances: misoprostol and diclofenac. Misoprostol and diclofenac dose per sample were as follows: 608.8 ng (sample 1), 708.4 ng (sample 2), 618.8 ng (sample 3) and 67.7 mg (sample 1), 65.3 mg (sample 2) 67.3 mg (sample 3), respectively. A simple, precise and reliable method can be applied for routine examinations in terms of clinical and forensic toxicology examinations as well as in quality control of drugs for pharmaceutical purposes (original drugs and counterfeit medications).

Dynamic profiling of intact glucosinolates in radish by combining UHPLC-HRMS/MS and UHPLC-QqQ-MS/MS.

Glucosinolates (GSLs) and their degradation products in radish confer plant defense, promote human health, and generate pungent flavor. However, the intact GSLs in radish have not been investigated comprehensively yet. Here, an accurate qualitative and quantitative analyses of 15 intact GSLs from radish, including four major GSLs of glucoraphasatin (GRH), glucoerucin (GER), glucoraphenin (GRE), and 4-methoxyglucobrassicin (4MGBS), were conducted using UHPLC-HRMS/MS in combination with UHPLC-QqQ-MS/MS. Simultaneously, three isomers of hexyl GSL, 3-methylpentyl GSL, and 4-methylpentyl GSL were identified in radish. The highest content of GSLs was up to 232.46 µmol/g DW at the 42 DAG stage in the 'SQY' taproot, with an approximately 184.49-fold increase compared to the lowest content in another sample. That the GSLs content in the taproots of two radishes fluctuated in a similar pattern throughout the five vegetative growth stages according to the metabolic profiling, whereas the GSLs content in the '55' leaf steadily decreased over the same period. Additionally, the proposed biosynthetic pathways of radish-specific GSLs were elucidated in this study. Our findings will provide an abundance of qualitative and quantitative data on intact GSLs, as well as a method for detecting GSLs, thus providing direction for the scientific progress and practical utilization of GSLs in radish.

Simultaneous determination of nine phenolic compounds in imitation wild Dendrobium officinale samples using ultrahigh-performance liquid chromatography-tandem mass spectrometry.

Dendrobium officinale Kimura et Migo (D. officinale), one of the nine everlasting types of grass, has gained increasing attention owing to its important roles in alternative medicines and drug discovery. Due to its natural resources being in danger of being extinct, imitation wild planting is becoming increasingly common. To assess the product's quality completely, an efficient ultrahigh performance liquid chromatography-triple quadrupole tandem mass spectrometry (UHPLC-QQQ-MS/MS) method was established to simultaneously quantify nine phenolic compounds in D. officinale samples. The extraction parameters, including solvent, solvent concentration, solid-liquid ratio, and extraction time, were systematically optimized with the single-factor test. The results demonstrated that extraction with a 1:200 solid-to-liquid ratio of 80% methanol for 1.5 h was the most efficient condition for the extraction of flavonoids. Satisfactory retention times and resolution of the nine analytes were acquired on the Thermo Scientific Hypersil GOLD column with multiple reaction monitoring in negative ion scanning mode. The method was validated to demonstrate its selectivity, linearity, precision, accuracy, and robustness. Thus, the verified UHPLC-QQQ-MS/MS method was successfully applied to the quantification of phenolic components present in D. officinale samples. The results indicated that the quantity and composition of phenolic components in D. officinale from various provenances were significantly different. This work provides a theoretical foundation for the cultivation and assessment of wild D. officinale quality.

Widely targeted metabolomics and HPLC analysis elaborated the quality formation of Yunnan pickled tea during the whole process at an industrial scale.

Yunnan pickled tea is produced from fresh tea-leaves through fixation, rolling, anaerobic fermentation and sun-drying. In this study, widely targeted metabolomics using UHPLC-QQQ-MS/MS and HPLC analysis were carried out to elaborate its quality formation during the whole process. Results confirmed the contribution of preliminary treatments and anaerobic fermentation to the quality formation. A total of 568 differential metabolites (VIP > 1.0, P < 0.05, FC > 1.50 or < 0.67) were screened through OPLS-DA. (-)-Epigallocatechin and (-)-epicatechin significantly (P < 0.05) increased from the hydrolyzation of ester catechins, such as (-)-epigallocatechin gallate and (-)-epicatechin gallate in anaerobic fermentation. Additionally, the anaerobic fermentation promoted vast accumulations of seven essential amino acids, four phenolic acids, three flavones and flavone glycosides, pelargonidin and pelargonidin glycosides, flavonoids and flavonoid glycosides (i.e. kaempferol, quercetin, taxifolin, apigenin, myricetin, luteolin and their glycosides) through relevant N-methylation, O-methylation, hydrolyzation, glycosylation and oxidation.

An ultra-sensitive UHPLC-QqQ-MS/MS method for determination of 54 benzodiazepines (pharmaceutical drugs, NPS and metabolites) and z-drugs in biological samples.

Benzodiazepines exhibit central nervous system depressive activity as well as sedative, hypnotic, and anticonvulsant properties, which enable to use them as medical treatment in anxiety, epilepsy, insomnia and alcohol withdrawal syndrome. However, from 2000s illegal benzodiazepine derivatives have started to emerge on illicit drug market as new psychoactive substances (NPSs) monitored in many countries. Analysis of both pharmaceutical drugs and NPSs from benzodiazepines group could be challenging, as usually very low concentrations need to be determined. Thus, an ultra-sensitive UHPLC-QqQ-MS/MS method was developed for simultaneous determination of 54 benzodiazepines (pharmaceutical drugs, NPS and their metabolites) and 3 z-drugs with one metabolite in biological fluid samples. The lower limit of quantification for most substances was 50 pg/mL, whereas for 17 substances as low as 10 pg/mL was achieved. Together with reduced sample volume to 100 µL it makes the developed method suitable for a sensitive multidrug toxicological analysis. Presented method was applied in routine toxicological practice as well as for the determination of benzodiazepines, z-drugs and their metabolites in 25 authentic biological fluids (blood, urine, vitreous humor and bile), both antemortem and postmortem. 19 different compounds, including benzodiazepines, their metabolites and z-drugs were determined. Antemortem blood concentrations were within 0.2-114.5 ng/mL, whereas concentrations in antemortem urine samples were 0.03-102.6 ng/mL. In postmortem specimens, concentrations ranged within 0.2-473.2 ng/mL, 0.5-94.1 ng/mL, 1.3-208.8 ng/mL and 41.5-42.0 ng/mL in blood, vitreous humor, urine and bile, respectively. The developed method is suitable for a forensic toxicology analysis, as well as clinical toxicology which is evidenced by the positive results of international proficiency tests.