Ultra-high Performance Liquid Chromatography-triple Quadrupole Research Articles

3α-Hydroxylup-20(29)-ene-23,28-dioic Acid as a Phytogenic Chemical Marker for Authenticating Schefflera octophylla (Lour.) Harms Monofloral Honey.

The monofloral honey from Schefflera octophylla (Lour.) Harms (MH-Sco) are of high economic value due to their rarity and potential medicinal benefits. However, the limited investigations on the relationship of phytogenic components between the plant S. octophylla (P-Sco) and MH-Sco have an impact on MH-Sco authentication. Herein, the tentative phytogenic markers of MH-Sco were screened by comparing the metabolites of MH-Sco obtained by ultrahigh-performance liquid chromatography-quadrupole time-of-flight tandem mass spectrometry (UHPLC-Q-TOF-MS/MS)-based untargeted metabolomics with the identified phytogenic chemicals from P-Sco. Combined with the mass and NMR spectral information, 3α-hydroxylup-20(29)-ene-23,28-dioic acid (HLEDA) was finally identified as the phytogenic marker of MH-Sco. A targeted ultrahigh-performance liquid chromatography-triple quadrupole mass spectrometry (UHPLC-QqQ-MS/MS)-based method was established and validated based on the purified monomer standard to measure HLEDA levels in honey samples. HLEDA determined in MH-Sco was with the content from 0.303 to 0.440 mg/kg, while HLEDA was absent in honey samples from other botanical origins, indicating the reliability of HLEDA as a chemical marker in MH-Sco authentication. This study provides the theoretical basis and industry guidance for honey quality control for commercial consumption.

Unambiguous identification of γ-aminobutyric acid adducts as novel plant biomarkers and their ultra-sensitive detection by UPLC–MS/MS for retrospective analyses of nitrogen mustards exposure

Plants are widely existing in the environments and have been considered as potential sentinel species of toxic chemicals′ exposure. In this study, the deadly toxic chemicals of three nitrogen mustards (NMs, including NH1, NH2 and NH3) were selected as the investigated targets. First, the reactivities of common endogenous plant components with NMs were examined in vitro. Then, the model plant Nicotiana benthamiana Domin was exposed to NMs. Three γ-aminobutyric acid–nitrogen mustard adducts (GABA–NMs) were identified in the living plant by high resolution mass spectrometry and comparison with the synthesized references. A sensitive detection method with the limits of quantification of 0.0500 ng mL−1 was developed using ultrahigh performance liquid chromatography–triple quadrupole mass spectrometry. The GABA–NMs could be detected after 120 days of the exposure and even in the dead leaves without obvious decrease. Furthermore, 20 different plant species grown in diverse climate zones were exposed to HN1, and the adduct of GABA–HN1 was identified in all the leaves. The results showed the good universality and specificity of GABA–NMs as plant biomarkers for NMs exposure. This work provides a new approach for the pollution investigation of toxic chemicals through analysing biomarkers in plant materials.

Simultaneous determination of seven artemisinin-related compounds in Artemisia annua by UPLC-QQQ-MS/MS

A variety of compounds in Artemisia annua were simultaneously determined to evaluate the quality of A. annua from multiple perspectives. A method based on ultra-high performance liquid chromatography-triple quadrupole tandem mass spectrometry(UPLC-QQQ-MS/MS) was established for the simultaneous determination of seven compounds: amorpha-4,11-diene, artemisinic aldehyde, dihydroartemisinic acid, artemisinic acid, artemisinin B, artemisitene, and artemisinin, in A. annua. The content of the seven compounds in different tissues(roots, stems, leaves, and lateral branches) of A. annua were compared. The roots, stems, leaves, and lateral branches of four-month-old A. annua were collected and the content of seven artemisinin-related compounds in different tissues was determined. A multi-reaction monitoring(MRM) acquisition mode of UPLC-QQQ-MS/MS was used, with a positive ion mode of atmospheric pressure chemical ion source(APCI). Chromatographic separation was achieved on an Eclipse Plus RRHD C_(18) column(2.1 mm×50 mm, 1.8 μm). The gradient elution was performed with the mobile phase consisted of formic acid(0.1%)-ammonium formate(5 mmol·L~(-1))(A) and the methanol(B) gradient program of 0-8 min, 55%-100% B, 8-11 min, 100% B, and equilibrium for 3 min, the flow rate of 0.6 mL·min~(-1), the column temperature of 40 ℃, the injection volume of 5 μL, and the detection time of 8 min. Through methodological investigation, a method based on UPLC-QQQ-MS/MS was established for the simultaneous quantitative determination of seven representative compounds involved in the biosynthesis of artemisinin. The content of artemisinin in A. annua was higher than that of artemisinin B, and the content of artemisinin and dihydroartemisinic acid were high in all the tissues of A. annua. The content of the seven compounds varied considerably in different tissues, with the highest levels in the leaves and neither artemisinene nor artemisinic aldehyde was detected in the roots. In this study, a quantitative method based on UPLC-QQQ-MS/MS for the simultaneous determination of seven representative compounds involved in the biosynthesis of artemisinin was established, which was accurate, sensitive, and highly efficient, and can be used for determining the content of artemisinin-related compounds in A. annua, breeding new varieties, and controlling the quality of Chinese medicinal materials.

Determination of 145 pharmaceuticals and personal care products in eleven categories in water by ultra-high performance liquid chromatography-triple quadrupole mass spectrometry

Pharmaceuticals and personal care products (PPCPs) are emerging contaminants frequently detected in aquatic environments at trace levels. These chemicals have diverse structures and physicochemical properties and includes pharmaceuticals like antibiotics, antihypertensive drugs, antiviral drugs, and psychotropic drugs that are widely used in large quantities worldwide. Considering the large number of pharmaceuticals currently in usage, it is crucial to establish a priority list of PPCPs that should be monitored and/or treated first. An accurate understanding of the occurrence and levels of PPCPs in aquatic environments is essential for providing objective materials for monitoring these emerging contaminants. Therefore, accurate, efficient, sensitive, and high-throughput screening techniques need to be established for determining and quantifying PPCPs. This study developed a method for the determination of 145 PPCPs (grouped into eleven categories: antibiotics, antihypertensive drugs, antidiabetic drugs, antiviral drugs, β-receptor agonists, nitroimidazoles, H2 receptor antagonists, psychotropic drugs, hypolipidemic drugs, non-steroidal anti-inflammatory drugs, and others) in water. The method was based on large volume direct injection without sample enrichment and cleanup and used ultra-high performance liquid chromatography-triple quadrupole mass spectrometry (UHPLC-MS/MS). Water samples were collected and filtered through a 0.22-μm regenerated cellulose (RC) filter membrane. Subsequently, Na2EDTA was added to the samples to adjust their pH to 6.0-8.0. Internal standards were mixed with the solutions, and because of the addition of Na2EDTA, the interference of metal ions could be eliminated in the determination of compounds, especially for tetracycline and quinolone antibiotics. Among the six filter membranes tested in this study (PES, PFTE-Q, PFTE, MCE, GHP, and RC), RC filter membranes were screened for water sample filtration. The UHPLC-MS/MS parameters were optimized by comparing the results of various mobile phases, as well as by establishing the best instrumental conditions. The 145 PPCPs were separated using an Phenomenex Kinetex C18 column (50 mm×3 mm, 2.6 μm) via gradient elution. The mobile phases were 0.1% (v/v) formic acid aqueous solution containing 5 mmol/L ammonium formate and acetonitrile for positive ion modes, 5 mmol/L aqueous solutions of ammonium formate and acetonitrile for negative ion modes. The samples were quantified using the scheduled multiple reaction monitoring (scheduled-MRM) mode with electrospray ionization in positive and negative ion modes. A standard internal calibration procedure was used to calculate contents of sample. The established method was systematically verified, and it demonstrated a good linear relationship. The average recoveries of the 145 PPCPs at the three spiked levels were in the range of 80.4%-128% with relative standard deviations (RSDs, n=6) of 0.6%-15.6%. The method detection limits (MDLs) ranged from 0.015 to 5.515 ng/L. Finally, the optimization method was applied to analyze the 145 PPCPs in 11 surface water samples and 6 drinking water samples. Overall, 93 (64%) out of the 145 analytes were detected. The total contents of the PPCPs in surface water samples ranged from 276.9 to 2705.7 ng/L. The detection frequencies of antidiabetic, antiviral, and psychotropic drugs were 100%. The total contents of the PPCPs in drinking water samples ranged from 140.5 to 211.5 ng/L, and antibiotics, antidiabetic drugs, and antiviral drugs comprised the largest proportion of analytes (by mass concentration) in drinking water samples. Our method exhibited high analytical speed and high sensitivity. It is thus suitable for the trace analysis and determination of the 145 PPCPs in environmental water and showed improved detection efficiency for PPCPs in water, indicating that it has a high potential for practical applications. This study can extend technical support for further pollution-level analysis of PPCPs in water and provide an objective basis for environmental management.

Determination of 118 pesticide residues in urine by ultra-high performance liquid chromatography-tandem mass spectrometry

Pesticide residues in food and their hazardous effects have attracted much attention given the increased and widespread use of pesticides. The long-term consumption of food containing pesticide residues is an important pathway for the gradual accumulation of pesticides in the human body. Urine is often monitored as a biological sample for low-dose exposure to pesticides, and urine collection is a relatively convenient sampling technique in general population research. In order to effectively monitor residual levels of multiple pesticides in human urine and provide an important technological approach for health risk assessment, a rapid screening and confirmatory detection method for 118 pesticides in urine was established using QuEChERS method as a pretreatment combined with ultra-high performance liquid chromatography-triple quadrupole mass spectrometry (UHPLC-MS/MS). The 118 pesticides analyzed included organophosphorus, carbamate, neonicotinoid, and strobilurin fungicides and other widely used pesticides. Following systematic optimization of the pretreatment process, LC separation conditions, and MS/MS parameters, 118 pesticides were extracted from urine samples and analyzed within 2 h. In brief, the target analytes in 5 mL urine samples were extracted with 10 mL of acetonitrile and added with 5 g of anhydrous MgSO4 and 1 g of NaCl as water-removal and salting-out agents, respectively. After centrifugation, 6 mL of the supernatant was cleaned using the QuEChERS method with 300 mg of C18, 300 mg of primary secondary amine (PSA) and 900 mg of anhydrous MgSO4 as the purification adsorbent. After nitrogen blowing and solubilization, the 118 target analytes were separated on a ZORBAX Eclipse Plus C18 analytical chromatographic column (100 mm×2.1 mm, 1.8 μm) with gradient elution using (A) 0.01% formic acid aqueous solution (containing 2 mmol/L ammonium formate) and (B) 0.01% formic acid methanol solution (containing 2 mmol/L ammonium formate) as mobile phases. The gradient elution program was as follows: 0-0.5 min, 5%B; 0.5-1.5 min, 5%B-20%B; 1.5-2.5 min, 20%B-50%B; 2.5-8.0 min, 50%B-80%B; 8.0-9.0 min, 80%B-98%B; 9.0-11.0 min, 98%B; 11.0-11.5 min, 98%B-5%B; 11.5-15.0 min, 5%B. The analytes were then determined by UHPLC-MS/MS with positive/negative ion switching in dynamic multiple-reaction monitoring mode and quantified using the external standard method. The results indicated that the proposed method can determine 118 pesticides in urine simultaneously and rapidly. The limits of detection and limits of quantification were 0.10 and 0.50 μg/L, respectively, and the matrix effects were less than 20%for all targeted compounds. The recoveries of the 118 pesticides in urine were between 70.2% and 104% at three spiked levels of 0.50, 1.00, and 5.00 μg/L, and the relative standard deviations ranged from 2.8% to 9.3%. The method was applied to 10 actual urine samples, and the results revealed the presence of six pesticides, including thiamethoxam, clothianidin, acetamiprid, dinotefuran, isoproturon, and dimethomorph, with contents ranging from <LOQ to 3.65 μg/L. Thiamethoxam and clothianidin were detected most often in the samples, and their concentrations were highly correlated with the current use of these pesticides in edible agricultural products. The proposed method has the characteristics of high efficiency, sensitivity, and accuracy and is suitable for the rapid screening and determination of the 118 pesticide residues in human urine samples.

Quality evaluation of Angelica Sinensis Radix dispensing granules by integrating microvascular activity and chemical analysis

Ethnopharmacological relevanceAs a new form of crude slices of traditional Chinese medicine (TCM), traditional Chinese medicine dispensing granules (TCMDGs) have been used for clinical formula. It is necessary to evaluate whether the chemical composition and biological activity are consistent among the different batches. Angelica Sinensis Radix (ASR), the root of Angelica sinensis (Oliv.) Diels, is one of the most frequently used medicinal materials in gynecology, senile and cardiovascular diseases. In this paper, the quality of TCMDGs is examined taking the Angelica Sinensis Radix dispensing granules (ASRDGs) as a typical case. Aim of the studyIn this study, integrating bioequivalence and chemical analysis was used to evaluate the quality of dispensing granules taking ASRDGs as a typical case. Materials and methodsAccording to the clinical efficacy of ASR, the intestinal absorption liquid of ASRDGs (IAL-ASRDGs) in 15 batches prepared by the everted gut sac (EGS) method was used to evaluate its effects of vasodilatation on isolated vascular rings. Then, the chemical profiling analysis of IAL-ASRDGs from the 15 batches was carried out by ultra-high performance liquid chromatography coupled with quadrupole-time of flight mass spectrometry (UPLC-Q-TOF-MS). The components in IAL-ASRDGs were identified using mass spectrometry data and the obtained reference standards. Pearson correlation analysis was further performed for the selection of quality control markers based on the extracted ion chromatograms of the identified compounds and vasodilatory activities of different batches of IAL-ASRDGs. Moreover, the selected chemical components in ASRDGs were further verified by vasodilatory activities and quantitatively analyzed by ultra-high performance liquid chromatography-triple quadrupole mass spectrometry (UPLC-QQQ-MS). ResultsThe IAL-ASRDGs showed favorable vasodilatory activities. There were significant differences among the 15 batches. The ranges of maximum vasodilation rate (%) and EC50 were 69.33 ± 7.16 to 19.52 ± 5.05 and 0.07–25.58 g raw materials/mL, respectively. A total of 46 compounds in IAL-ASRDGs were identified based on accurate mass measurements, fragmentation behavior and the reference standards. Among them, 8 compounds including butylidenephthalide, butylphthalide and senkyunolide A showed positive correlation with vasodilatory effect of IAL-ASRDGs. The 8 compounds were further verified, showing obvious vasodilatory activities. The content of the 8 compounds also showed some differences in 15 batches of ASRDGs. Among them, the content of ferulic acid, senkyunolide I and senkyunolide H varied the most in different batches of ASRDGs. By Pearson correlation analysis, the total content of senkyunolides (senkyunolide A, senkyunolide I and senkyunolide H) in ASRDGs was found to reflect the vasodilatory activity of ASRDGs mostly. ConclusionThis study provides new strategies for the quality assessment by bioequivalence and explore the chemical quality control markers for ASRDGs.

Effect of BBM gene on callus growth and ginsenoside content in Panax quinquefolius

Baby Boom(BBM) gene is a key regulatory factor in embryonic development and regeneration, cell proliferation, callus growth, and differentiation promotion. Since the genetic transformation system of Panax quinquefolius is unstable with low efficiency and long period, this study attempted to transfer BBM gene of Zea mays to P. quinquefolius callus by gene gunship to investigate its effect on the callus growth and ginsenoside content, laying a foundation for establishing efficient genetic transformation system of P. quinquefolius. Four transgenic callus of P. quinquefolius with different transformation events were obtained by screening for glufosinate ammonium resistance and molecular identification by PCR. The growth state and growth rate of wild-type and transgenic callus were compared in the same growth period. The content of ginsenoside in transgenic callus was determined by ultra-high performance liquid chromatography-triple quadrupole mass spectrometry(UPLC-MS/MS). The results showed that transgenic callus growth rate was significantly higher than that of wild-type callus. In addition, the content of ginsenoside Rb_1, Rg_1, Ro, and Re was significantly higher than that in wild-type callus. The paper preliminarily proved the function of BBM gene in promoting growth rate and increasing ginsenoside content, which provided a scientific basis to establish a stable and efficient genetic transformation system for Panax plants in the future.

Multifunctionalized Covalent Organic Frameworks for Broad-Spectrum Extraction and Ultrasensitive Analysis of Per- and Polyfluoroalkyl Substances

The contamination of surface and ground water by per- and polyfluoroalkyl substances (PFASs) has become a growing concern, and the structural diversity of PFASs is the major challenge for their ubiquitous applications. Strategies for monitoring coexistent anionic, cationic, and zwitterionic PFASs even at trace levels in aquatic environments are urgently demanded for effective pollution control. Herein, novel amide group and perfluoroalkyl chain-functionalized covalent organic frameworks (COFs) named COF-NH-CO-F9 are successfully synthesized and used for highly efficient extraction of broad-spectrum PFASs, attributing to their unique structure and the multifunctional groups. Under the optimal conditions, a simple and high-sensitivity method is established to quantify 14 PFASs including anionic, cationic, and zwitterionic species by coupling solid-phase microextraction (SPME) with ultrahigh-performance liquid chromatography-triple quadrupole mass spectrometry (UHPLC-MS/MS) for the first time. The established method displays high enrichment factors (EFs) of 66-160, ultrahigh sensitivity with low limits of detection (LODs) of 0.0035-0.18 ng L-1, a wide linearity of 0.1-2000 ng L-1 with correlation coefficient (R2) ≥0.9925, and satisfactory precision with relative standard deviations (RSDs) ≤11.2%. The excellent performance is validated in real water samples with recoveries of 77.1-108% and RSDs ≤11.4%. This work highlights the potential of rational design of COFs with the desired structure and functionality for the broad-spectrum enrichment and ultrasensitive determination of PFASs in real applications.

Discovery of the material basis of Jiuwei Xifeng granules using pharmaco-chemistry and pharmacokinetics

Ethnopharmacological relevanceJiuwei Xifeng granules (JWXF) is primarily used for the treatment of Tourette syndrome (TS) with kidney-Yin deficiency and internal stirring of liver wind. However, few studies have focused on this issue. Aim of the studyThis study aimed to clarify chemical composition of JWXF using in vitro and in vivo pharmaco-chemistry and to provide a basis for the clinical use of JWXF using a strategy of pharmacokinetics. Materials and methodsIn this study, the chemical constituents and in vivo metabolism of JWXF were evaluated using high performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (HPLC-Q-TOF-MS/MS), and the time-dependent processes of the three main components in rats were detected using ultra-high performance liquid chromatography-triple quadrupole mass spectrometry (UPLC-QQQ-MS/MS). ResultsA total of 75 constituents were identified, including 22 alkaloids, 21 terpenes, 15 organic acids and their derivatives, and 17 other compounds. After administration, 12 compounds were identified in rat plasma, including 11 prototypes and one metabolite. Pharmacokinetic analysis showed that the effects of gentiopicroside, gastrodin, and sweroside in rats were dose-dependent when the dose of JWXF was 1–4 g/kg. They were rapidly absorbed and did not accumulate in the plasma after 7-day continuous intragastric administration. ConclusionsJWXF consists of 75 components, including alkaloids, terpenes, and organic acids. The three main compounds, gastrodin, gentiopicroside, and sweroside, undergo rapid absorption, elimination, and dose-dependent pharmacokinetics.

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.

Identification of Nutritional Ingredients and Medicinal Components of Hawk Tea and Insect Tea Using Widely Targeted Secondary Metabolomics

In this study, the metabolites in insect tea and hawk tea were analyzed by ultra-high performance liquid chromatography–triple quadrupole mass spectrometry. We found 49 metabolites in insect tea and hawk tea that can be used as key active components in traditional Chinese medicine, as well as 98 metabolites that can be used as active components of pharmaceutical preparations for the treatment of cancers, hypertension, cardiovascular diseases, etc. Comparative analysis found that insect tea and hawk tea had significant differences in their metabolic profiles. Insect tea contains more metabolites beneficial to human health than hawk tea; insect tea also has higher antioxidant activity in vitro than hawk tea. The results of this study will contribute to the development of new health foods using insect tea.

Pseudotargeted metabolomics analysis of pine pollen intervention in the liver of premature ovarian failure rats

Premature ovarian failure (POF) is a prevalent gynecological disease. In traditional Chinese medicine, it is believed that POF is directly related to abnormal function of the liver and kidneys. As such, regulation of the liver metabolism through the use of medicinal and edible substances is important for the treatment of POF. Pine pollen, a traditional Chinese medicinal and edible pollen variety, contains various active substances, such as sex hormones and phytohormones, which have been used to inhibit inflammation, regulate the immune system, and protect reproductive tissues. Using ultra-high performance liquid chromatography-triple quadrupole mass spectrometry (UHPLC-MS/MS), this study examined the influence of pine pollen on the liver metabolome of cyclophosphamide-induced POF model Sprague Dawley (SD) rats. The variations in the metabolites present in the liver tissue of control SD rats, model SD rats, and SD rats treated with various doses of pine pollen or estrogen were analyzed using principal component analysis (PCA) in combination with orthogonal partial least squares discriminant analysis (OPLS-DA) and other multivariate statistical methods to reveal the mechanism of pine pollen intervention in the livers of POF SD rats. An animal model experiment was conducted using six groups of ten-week-old rats. Cyclophosphamide was administered intraperitoneally to the model group and four intervention groups at a dosage of 60 mg/kg for 1 d followed by a dosage of 10 mg/kg for 14 d. Within the following four weeks, each of the four intervention groups received the intragastric administration of 0.1, 0.5, or 1.5 g/kg bodyweight (BW) of pine pollen, or 0.075 g/kg BW of conjugated estrogens (positive control). Equal quantities of normal saline were administered to the control and cyclophosphamide-treated model groups. Subsequently, the rat livers were subject to pseudotargeted metabolomics, and a total of 687 liver metabolites were discovered using both positive and negative ions. The metabolites differing in content were screened using the t-test (p<0.05) and the fold change (FC>2 or <0.5) in univariate analysis, and the variable importance in projection (VIP>1) in multivariate analysis. It was found that in comparison with the control group, the contents of 32 metabolites significantly increased, while those of 28 metabolites significantly decreased in the model group. The majority of these metabolites were involved α-linolenic acid metabolism, vitamin B6 metabolism, and purine metabolism, along with the lysine degradation and glycolysis/gluconeogenesis metabolic pathways. Compared with the cyclophosphamide-induced model group, the estrogen group exhibited increased levels of 47 metabolites and decreased levels of 29 metabolites, wherein 34 metabolites were restored to the levels found in the control group. These metabolites mainly involved the vitamin B6, lysine, glycolysis/gluconeogenesis, arginine and proline, and cysteine and methionine metabolic pathways. In the low/medium/high-dose pine pollen groups, the contents of 34/32/34 metabolites increased, the contents of 30/37/24 metabolites decreased, and the contents of 47/38/34 metabolites were restored to the levels found in the control group, respectively. These metabolites were mainly involved in vitamin B6 metabolism, purine metabolism, and the glycolysis/gluconeogenesis metabolic pathway. These results therefore indicate that the restoring effect of pine pollen is equivalent or superior to that of conjugated estrogen. Additionally, based on the known metabolic pathways, it appears that when estrogen interferes with the liver metabolism, the key metabolic pathways that become affected are the arginine and proline metabolism and cysteine and methionine metabolism pathways. In contrast, pine pollen intervention affected existing metabolic pathways that were known to be disordered by cyclophosphamide. The use of pine pollen may therefore restore the levels of many metabolites. It should be noted that 23 overlaps exist between the estrogen-restored metabolites and the pine pollen-restored metabolites, including a variety of acylcarnitines, such as ACar 10∶0. As a result, pine pollen extract may be able to normalize the liver metabolic abnormalities induced by POF. This study therefore establishes a theoretical reference for the development of functional applications for pine pollen and for the treatment of POF.

Quantification and purification of procyanidin B1 from food byproducts.

The flavan-3-ols of 10 primarily plant food byproducts, including MuscatHamburg grape seed, hawthorn sarcocarp, litchi pericarp, cocoa bean, peanut skin, lotus seedpod, Xinyang Maojian green tea, Cinnamomi cortex, Sargentodoxa cuneata stem, and Cyperus esculentus, leaves were analyzed. Ultrahigh-performance liquid chromatography-triple quadrupole mass spectrometry was used for the analysis. Cyperus esculentus leaves contained a high amount of procyanidin B1 (198.9mg/100g), second only to MuscatHamburg grape seed (292.9mg/100g). Unlike grape seed that contained several procyanidin B isomers with very similar retention times, C. esculentus leaves contained primarily procyanidin B1 with few isomers. Procyanidin B1 was enriched in the ethyl acetate fraction of a 70% methanol extract of C. esculentus leaves and purified at 95% purity by two runs of open column chromatography. Direct chromatography of the plant extract on octadecylsilane and Sephadex LH20 open columns improved the yield of the resultant leaf procyanidin B1 (95% purity) to 0.21‰. The present research demonstrated that the leaves of C. esculentus, byproducts of tigernut, are ideal plant sources for isolating and providing high-purity procyanidin B1. PRACTICAL APPLICATION: Procyanidin B1 has a broad range of health benefits. Cyperus esculentus is cultivated in many countries with nearly 6190 square hectares (hm2 ) in the Spanish Mediterranean region in 2020-2021 and over 16,700 hm2 in China in recent years, primarily for its tubers. The byproducts, the leaves of C. esculentus, contain high levels of procyanidin B1, with few isomers that interfere with its isolation and purification. Thus, the leaves of this plant provide a viable source for preparing high-purity procyanidin B1.

Residue behavior and risk assessment of pyraclostrobin and tebuconazole in peppers under different growing conditions.

This study evaluates the residue behavior and risks of pyraclostrobin and tebuconazole in peppers. An analytical method for the simultaneous determination of the concentration of these fungicides in peppers was developed using ultra-high performance liquid chromatography-triple quadrupole mass spectrometry. Pepper samples were extracted with acetonitrile and cleaned with primary secondary amine and graphitized carbon black. The average recoveries of pyraclostrobin and tebuconazole under three fortification levels were 86.7-101.4% and 81.7-104.4%, with relative standard deviations of 4.0-7.2% and 3.8-10.9%, respectively. The limit of quantification of both fungicides in peppers was 0.01mg/kg. The terminal residue trial of 30% pyraclostrobin and tebuconazole suspension concentrate was investigated for samples cultivated in open fields and greenhouses. The results showed that the terminal residues of pyraclostrobin and tebuconazole in peppers were lower than the maximum residue limits established by GB 2763-2021 (0.5mg/kg for pyraclostrobin and 2mg/kg for tebuconazole). The results of a statistical t-test indicated that there was no significant difference between samples grown in open fields and greenhouses. According to the international estimate of short-term intake (IESTI) calculation model, provided by the Joint FAO/WHO Meeting on Pesticide Residues, the acute dietary exposure risk of both fungicides in peppers was acceptable for the general population, with an IESTI of 0-3% and 0-5% of the acute reference dose for pyraclostrobin and tebuconazole, respectively.

Toxicokinetics of Two Oxathiapiprolin Enantiomers in Rats and Their Stereoselective Interaction with Oxysterol Binding Protein.

Oxathiapiprolin is a chiral fungicide, and it can affect the metabolism of the cholesterol compounds by inhibiting oxysterol binding protein (OSBP) to exert its fungicidal effect. The application of oxathiapiprolin in agriculture is widespread, and its residue in the environment is a threat to both human and animal health. The two oxathiapiprolin enantiomers differ in their fungicidal activity, biotoxicity, and degradation by environmental forces. However, their biotoxicity has not been reported in animals. The toxicokinetics of a pesticide should be a crucial component for the evaluation of its toxicity in vivo. In this study, we investigated the absorption, bioavailability, tissue distribution, and excretion of the two oxathiapiprolin enantiomers in rats to verify their toxicokinetic process in animals. An ultrahigh-performance liquid chromatography triple quadrupole tandem mass spectrometry (UHPLC-QQQ/MS) method was established to quantify the two oxathiapiprolin enantiomers in vivo. The two oxathiapiprolin enantiomers were found to have approximately the same absorption rate and bioavailability, and both were excreted mainly in the feces. The half-life of R-(-)-oxathiapiprolin was nearly twice that of S-(+)-oxathiapiprolin. R-(-)-oxathiapiprolin also had greater distribution than S-(+)-oxathiapiprolin in the liver, lungs, heart, spleen, kidneys, stomach, large intestine, small intestine, brain, and pancreas, supporting the notion that R-(-)-oxathiapiprolin could better bind with OSBP. The stereoselectivity of S-(+)-oxathiapiprolin in these tissues may be responsible for it being readily metabolized in vivo. The molecular docking technique was subsequently used to verify the more superior binding between R-(-)-oxathiapiprolin and OSBP compared with the binding between S-(+)-oxathiapiprolin and OSBP. The findings of this study could provide more reliable data for determining the toxicokinetics of a single enantiomer of oxathiapiprolin in animals, thereby providing some theoretical basis for its subsequent toxicological study.

Serum lipidomics profiles reveal potential lipid markers for prediabetes and type 2 diabetes in patients from multiple communities.

Dyslipidemia is a hallmark of diabetes mellitus (DM). However, specific lipid molecules closely associated with the initiation and progression of diabetes remain unclear. We used a pseudotargeted lipidomics approach to evaluate the complex lipid changes that occurred long before the diagnosis of type 2 diabetes mellitus (T2DM) and to identify novel lipid markers for screening prediabetes mellitus (PreDM) and T2DM in patients from multiple communities. Four hundred and eighty-one subjects consisting of T2DM, three subtypes of PreDM, and normal controls (NC) were enrolled as discovery cohort. Serum lipidomic profiles of 481 subjects were analyzed using an ultrahigh performance liquid chromatography-triple quadrupole mass spectrometry (UHPLC-QqQ-MS)-based pseudotargeted lipidomics method. The differential lipid molecules were further validated in an independent case-control study consisting of 150 PreDM, 234 T2DM and 94 NC. Multivariate discriminative analyses show that lipidomics data have considerable potential for identifying lipidome differences among T2DM, subtypes of PreDM and NC. Statistical associations of lipid (sub)species display significant variations in 11 lipid (sub)species levels for T2DM and distinctive differences in 8 lipid (sub)species levels between prediabetic and normoglycemic individuals, with further differences in 8 lipid (sub)species levels among subtypes of PreDM. Adjusted for sex, age and BMI, only two lipid (sub)species of fatty acid (FA) and phosphatidylcholine (PC) were associated at p< 0.05 for PreDM (all) and subtypes of PreDM. The defined lipid markers not only significantly improve the diagnostic accuracy of PreDM and T2DM but also effectively evaluating the risk of developing into each subtype of PreDM and T2DM when addition of age, sex, BMI, and FPG, respectively. Our findings improve insights into the lipid metabolic complexity and interindividual variations among subtypes of PreDM and T2DM, beyond the well-known differences in dyslipidemia in clinic.

Simultaneous quantification of multiple amino acid adducts from sulfur mustard-modified human serum albumin in plasma at trace exposure levels by ultra-high performance liquid chromatography–triple quadrupole mass spectrometry after propionyl derivatization

Sulfur mustard (HD) is a highly toxic vesicant and is prohibited by the Organisation for the Prohibition of Chemical Weapons (OPCW). HD can modify human serum albumin (HSA) to generate hydroxyethylthioethyl (HETE) adducts, which could be utilized as biomarkers for verifying HD exposure in forensic analysis. Here, five amino acid adducts generated from pronase digestion of HD-exposed human serum albumin (HD–HSA) in plasma were selected as biomarkers to retrospectively detect HD exposure. HD–HSA was precipitated from plasma with acetone, digested by pronase, derivatized with propionic anhydride (PA), and analysed with ultra-high performance liquid chromatography coupled with triple quadrupole mass spectrometry (UHPLC–TQ MS). The limits of detection (LODs) and limits of quantification (LOQs) of the HD exposure concentrations were evaluated as 1.00 ng/mL at S/N≥3 and 3.00 ng/mL at S/N≥10, respectively, which are approximately 60 times lower than those of the reported method. The approach shows good linearity (R2≥0.997) from 3.00 ng/mL to 10.0 µg/mL of HD-exposed human plasma with satisfactory precision and accuracy. The developed approach was applied to analysing samples from the 6th OPCW Biomedical Proficiency Test (BioPT). The study showed that the developed approach was also suitable for analysing human plasma samples that were exposed to six of HD analogues, which were common impurities in sulfur mustard mixtures. Moreover, the method was successfully applied to plasma from other species, including rabbits, rats and cattle. This study provides a reliable and sensitive tool for the retrospective detection of vesicants exposure based on multiple biomarkers.

Systematic Qualitative and Quantitative Analyses of Wenxin Granule via Ultra-High Performance Liquid Chromatography Coupled with Ion Mobility Quadrupole Time-of-Flight Mass Spectrometry and Triple Quadrupole-Linear Ion Trap Mass Spectrometry.

Wenxin granule (WXG) is a popular traditional Chinese medicine (TCM) preparation for the treatment of arrhythmia disease. Potent analytical technologies are needed to elucidate its chemical composition and assess the quality differences among multibatch samples. In this work, both a multicomponent characterization and quantitative assay of WXG were conducted using two liquid chromatography–mass spectrometry (LC-MS) approaches. An ultra-high performance liquid chromatography–ion mobility quadrupole time-of-flight mass spectrometry (UHPLC/IM-QTOF-MS) approach combined with intelligent peak annotation workflows was developed to characterize the multicomponents of WXG. A hybrid scan approach enabling alternative data-independent and data-dependent acquisitions was established. We characterized 205 components, including 92 ginsenosides, 53 steroidal saponins, 14 alkaloids, and 46 others. Moreover, an optimized scheduled multiple reaction monitoring (sMRM) method was elaborated, targeting 24 compounds of WXG via ultra-high performance liquid chromatography–triple quadrupole linear ion trap mass spectrometry (UHPLC/QTrap-MS), which was validated based on its selectivity, precision, stability, repeatability, linearity, sensitivity, recovery, and matrix effect. By applying this method to 27 batches of WXG samples, the content variations of multiple markers from Notoginseng Radix et Rhizoma (21) and Codonopsis Radix (3) were depicted. Conclusively, we achieved the comprehensive multicomponent characterization and holistic quality assessment of WXG by targeting the non-volatile components.

Evaluation of Multicomponent Changes of Schisandra chinensis Fruits with Different Drying Process by UPLC-QQQ-MS-Based Targeted Metabolomics Analysis

Schisandra chinensis fruit is a famous tonic traditional Chinese medicine and healthy food. The chemical components of dried fruits may vary with the initial processing methods of fresh fruits. Therefore, in this study, the influence of drying was evaluated based on targeted metabolomics analysis. The ultrahigh performance liquid chromatography-triple quadrupole mass spectrometry (UPLC-QQQ-MS) was applied in determination of multicomponent including lignans, organic acids, and oligosaccharides. The fresh fruits of Schisandra chinensis were processed by natural drying in the sun, hot air drying and vacuum drying at different temperatures, and vacuum freeze drying, respectively. The contents of lignans, organic acids, and oligosaccharides in different fruit samples were quantified and statistically analyzed by partial least squares discriminant analysis (PLS-DA). The variation of totally 10 lignans, 5 organic acids, and 9 oligosaccharides with different drying process were compared. The results showed that the drying time of natural drying method was much longer than the other methods using instrument. For hot air and vacuum drying samples, most of lignans, organic acids, and oligosaccharides gradually decreased with the increase of temperature. And the appearance of fruit drying at higher temperature was dark red. At the same processing temperature, vacuum drying takes longer time than hot air drying. In addition, organic acid and oligosaccharide contents of vacuum freeze drying treated samples were higher than the other processed samples significantly. According to the targeted metabolomics analysis, the lignans, organic acids, and oligosaccharides markers were screened out for holistic quality evaluation of Schisandra chinensis fruits. In comprehensive consideration of multicomponent, drying time, fruit appearance and storage, and production feasibility, hot air drying at 50°C is more suitable for the process of Schisandra chinensis. The result is expected to provide a scientific basis for the selection of proper method for drying process of fresh Schisandra chinensis fruit.

Rapid and selective determination of 9 nitrosamines in biological samples using ultra-high performance liquid chromatography-triple quadrupole linear ion trap mass spectrometry.

A sensitive, selective and convenient method for the simultaneous determination of 9 nitrosamines (NAs) in biological samples was developed using isotope dilution ultra-high performance liquid chromatography-triple quadrupole linear ion trap mass spectrometry (UPLC-QTRAP-MS). Multiple reaction monitoring-information dependent acquisition-enhanced product ion (MRM-IDA-EPI) scan mode was performed to eliminate false positive results, and the whole detection procedure was characterized by less time consuming and simple sample preparation. 9 NAs were separated through a T3 column with the gradient elution of acetonitrile and water, and detected by UPLC-QTRAP-MS with an atmospheric pressure chemical ionization (APCI) source in the positive mode. The quantitative analysis was carried out via the isotope internal standard method with a matrix calibration curve. Under the optimized conditions, good linearity for the 9 NAs was achieved in the range of 0.2-20 μg L-1 with correlation coefficients (r) higher than ≥0.9991, and the limits of detection and limits of quantitation were 0.02-0.1 μg L-1 (S/N = 3) and 0.06-0.3 μg L-1 (S/N = 10), respectively. Satisfactory recoveries ranging from 79.4% to 108.0% were obtained, and the precision of the proposed method, indicated by the relative standard deviations (RSDs), was 2.3-12.9%. The matrix effect study showed that NDMA, NMOR and NMEA presented a matrix suppression effect, NDPHA displayed a matrix enhancement effect, and the matrix effects of the other 5 analytes could be ignored. Real application of the developed method in 13 urine and 24 plasma samples demonstrated that NDBA, NPIP and NPYR occurred in both urine and plasma samples with the concentration of 0.038-0.60 μg L-1, while other NAs were not detected. Such a method was sensitive and selective, and could be applied to the rapid qualitative and quantitative analysis of the 9 NAs in biological samples.