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Context As an inhibitor cytochrome P450 family 2 subfamily C polypeptide 8 (CYP2C8), quercetin is a naturally occurring flavonoid with its glycosides consumed at least 100 mg per day in food. However, it is still unknown whether quercetin and selexipag interact. Objective The study investigated the effect of quercetin on the pharmacokinetics of selexipag and ACT-333679 in beagles. Materials and methods The ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was used to investigate the pharmacokinetics of orally administered selexipag (2 mg/kg) with and without quercetin (2 mg/kg/day for 7 days) pre-treatment in beagles. The effect of quercetin on the pharmacokinetics of selexipag and its potential mechanism was studied through the pharmacokinetic parameters. Results The assay method was validated for selexipag and ACT-333679, and the lower limit of quantification for both was 1 ng/mL. The recovery and the matrix effect of selexipag were 84.5–91.58% and 94.98–99.67%, while for ACT-333679 were 81.21–93.90% and 93.17–99.23%. The UPLC-MS/MS method was sensitive, accurate and precise, and had been applied to the herb-drug interaction study of quercetin with selexipag and ACT-333679. Treatment with quercetin led to an increased in Cmax and AUC0–t of selexipag by about 43.08% and 26.92%, respectively. While the ACT-333679 was about 11.11% and 18.87%, respectively. Discussion and conclusion The study indicated that quercetin could inhibit the metabolism of selexipag and ACT-333679 when co-administration. Therefore, the clinical dose of selexipag should be used with caution when co-administered with foods high in quercetin.

Hui-jun Wang,1 Chun-yan Zhou,1 Yan-ding Su,1 Kai-feng Gou,1 Xiao-nan Geng,1 Xiang-jun Qiu1,2 1Department of Pharmacy, School of Basic Medical Sciences, Henan University of Science and Technology, Luoyang, 471023, People’s Republic of China; 2Functional Experiment Teaching Center, School of Basic Medical Sciences, Henan University of Science and Technology, Luoyang, 471023, People’s Republic of ChinaCorrespondence: Xiang-jun QiuFunctional Experiment Teaching Center, School of Basic Medical Sciences, Henan University of Science and Technology, 263 Kaiyuan Avenue, Luoyang, 471023, People’s Republic of ChinaTel +8613698882699Fax +86379-64830346Email lyxiangjun@126.comBackground: A new UPLC-MS/MS technique for the determination of ripretinib in beagle dog plasma was developed, and the pharmacokinetic effects of voriconazole and itraconazole on ripretinib in beagle dogs were studied.Methods: After extraction with ethyl acetate under alkaline conditions, ripretinib was detected using avapritinib as the internal standard (IS). The mobile phases were 0.1% formic acid-acetonitrile. The scanning method was multi-reaction monitoring using ESI+ source, and the ion pairs for ripretinib and IS were m/z 509.93→ 416.85 and 499.1→ 482.09, respectively. This animal experiment adopted a three period self-control experimental design. In the first period, ripretinib was orally administered to six beagle dogs at a dose of 5 mg/kg. In the second period, the same six beagle dogs were orally given itraconazole at a dose of 7 mg/kg, after 30 min, ripretinib was orally given. In the third period, voriconazole at a dose of 7 mg/kg was given orally, and then ripretinib was orally given. At different time points, the blood samples were collected. The concentration of ripretinib was detected, and the pharmacokinetic parameters of ripretinib were calculated.Results: Ripretinib had a good linear relationship in the range of 1– 1000 ng/mL. The precision, accuracy, recovery, matrix effect and stability met the requirements of the guiding principles. After erdafitinib combined with itraconazole, the Cmax and AUC0→t of ripretinib increased by 38.35% and 36.36%, respectively, and the t1/2 was prolonged to 7.53 h. After ripretinib combined with voriconazole, the Cmax and AUC0→t of ripretinib increased by 37.44% and 25.52%, respectively, and the t1/2 was prolonged to 7.33 h.Conclusion: A new and reliable UPLC-MS/MS technique was fully optimized and developed to detect the concentration of ripretinib in beagle dog plasma. Itraconazole and voriconazole could inhibit the metabolism of ripretinib in beagle dogs and increase the plasma exposure of ripretinib.Keywords: itraconazole, voriconazole, ripretinib, UPLC-MS/MS, pharmacokinetics, beagle dog

Meng-Jie Wang,1 Yu-Hang Zhao,2 Chen Fan,3 Ying-Jie Wang,3 Xin-Qi Wang,3 Xiang-Jun Qiu,3 Rui-Le Shen1 1Department of Neurology, The First Affiliated Hospital, and College of Clinical Medicine of Henan University of Science and Technology, Luoyang, Henan, 471003, People’s Republic of China; 2School of Medical Imaging, Hangzhou Medical College, Hangzhou, 310051, Zhejiang, People’s Republic of China; 3Department of Pharmacy, School of Basic Medical Sciences, Henan University of Science and Technology, Luoyang, 471023, Henan, People’s Republic of ChinaCorrespondence: Xiang-Jun QiuDepartment of Pharmacy, School of Basic Medical Sciences, Henan University of Science and Technology, Luoyang, 471023, Henan, People’s Republic of ChinaEmail lyxiangjun@126.comRui-Le ShenDepartment of Neurology, The First Affiliated Hospital, and College of Clinical Medicine of Henan University of Science and Technology, Luoyang, 471003, Henan, People’s Republic of ChinaEmail yfyshenruile@126.comBackground: Upadacitinib, a novel selective Janus kinase 1 (JAK1) inhibitor, has been recently approved by the US FDA for the treatment of adult patients with moderately to severely active rheumatoid arthritis (RA). An ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method for the quantitative analysis of upadacitinib in beagle dog plasma was developed and validated.Methods: Upadacitinib and fedratinib (internal standard, IS) were extracted with ethyl acetate under alkaline condition and then separated and detected. The chromatographic column was Waters Acquity UPLC BEH C18 column (2.1 mm × 50 mm, 1.7 μm), the mobile phase was acetonitrile and 0.1% formic acid in water with gradient elution procedure, and the flow rate was 0.40 mL/min. Under the positive ion mode, upadacitinib and IS were monitored by multiple reaction monitoring (MRM) as the following mass transition pairs: m/z 447.00 → 361.94 for upadacitinib and m/z 529.82 → 141.01 for IS.Results: In the concentration range of 1– 500 ng/mL, upadacitinib had good linearity, and the lower limit of quantification (LLOQ) was 1 ng/mL. The RSD of the intra- and inter-day precision was less than 10.03%, and the RE of accuracy was − 3.79% to 2.58%. The extraction recovery of upadacitinib was more than 80%, the matrix effect was around 100%, and upadacitinib was found to be stable.Conclusion: The novel optimized UPLC-MS/MS assay was an effective tool for the determination of upadacitinib and had been successfully applied to the pharmacokinetic study of upadacitinib in beagle dogs, and this method would also be used to study DDIs.Keywords: upadacitinib, UPLC-MS/MS, pharmacokinetic, beagle dog

For the simultaneous identification and quantification of five nitrofurans metabolites in farmed shrimp and fish, 3-amino-2-oxazolidinone (AOZ), 3-amino-5-morpholinomethyl-2-oxazolidinone (AMOZ), 1-aminohydantoine (AHD), semicarbazide (SEM), and 3,5-dinitrosalicylic acid hydrazide (DNSH), an accurate, precise, and specific method was developed. The mixture of water and methanol (60/40; v/v) was found to be the final optimised solvent for injection. The analytical run duration was 7 min, and the mobile phase included 2 mM methanol and ammonium formate. The new reference point for action (RPA) of 0.50 µg kg−1 as per EC/1871/2019 was taken into consideration and evaluated for the performance characteristics as per the CIR (EC)/2021/808 criteria. Specificity, relative retention time (≤0.25%) relative ion ratio (≤40%), linearity (0.25 to 2.0 µg kg−1), trueness (between 82.8 and 118.1%), repeatability (RSDr ≤14%), within lab reproducibility (RSDwr ≤16.9%), CCα (0.32–0.36 μg kg−1), ruggedness and relative matrix effect (≤14.26%) achieved acceptable values.

Infant milk formula was used as a model food to compare the sensitivity of thiobarbituric acid reactive substances (TBARS) and hydroperoxide methods to UPLC-MS/MS oxylipin analysis for detecting early lipid oxidation. Two different infant milk formulas were tested during 21 days of storage at 4°C. Formulas 1 and 2 contained canola oil and canola oil + 1% docosahexaenoic acid ethyl ester, respectively. Formulas were sampled up to 21 days of storage. Formula 2 had higher peroxide values than Formula 1 across all time points. However, no significant differences over time in TBARS and peroxide values in either formula were observed. Several oxylipins increased in both formulas starting on day 7 (linoleic acid and alpha-linolenic acid-derived oxylipins in Formula 1 and DHA-derived oxylipins in Formula 2). These results indicate that free oxylipins are effective in detecting early lipid oxidation and distinguishing between formulations containing different fatty acids. PRACTICAL APPLICATION: We have recently shown that primary oxidation products known as oxylipins can be measured in their free form by ultra-high pressure liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) to detect early lipid oxidation. However, a head-to-head comparison of the sensitivity of this approach to conventional spectrophotometric methods has not been evaluated. Our results indicate that free oxylipin measurements are better than conventional methods in detecting early lipid oxidation in milk infant formula distinguishing between different formulations.

In this study, UPLC-MS/MS was used to study the effects of smoking duration and temperature on the formation of heterocyclic amines (HAs) in smoke-processed meat patties. Four kinds of free HAs—including F-7,8-DiMeIQx; F-MeAαC; F-Harman and F-Norharman—and six kinds of protein-bound HAs—including B-AαC; B-7,8-DiMeIQx; B-Glu-p-1; B-MeAαC; B-Harman and B-Norharman—were detected and quantified. Among the free HAs, we observed a 23-fold content increase (p < 0.05), from 0–4 h (at 0 h and 4 h they were 4.24 ng·g−1 and 98.33 ng·g−1, respectively), and the content of the free HAs decreased to 78.80 ng·g−1, at 5 h. At the same time, the free HAs content increased from 53.52 ng·g−1, at 50 °C, to 127.16 ng·g−1, at 60 °C, and then decreased continuously. The content of the free HAs was the highest at 60 °C. For the protein-bound HAs, their content was found to generally decrease with the increase in smoking duration and temperature. However, at 5 h, the content of protein-bound HAs slightly increased to 984.2 ng·g−1. Meanwhile, at 90 °C, it increased to 1643.53 ng·g−1. Additionally, a total of 16 volatile organic compounds (VOCs) were found in all of the meat samples, of which 10 VOCs (one acid, three aldehydes and seven phenols) were significantly related to the formation of free HAs. These findings showed that all the different types of HAs were produced under low-temperature processing, which provided scientific insights into the potential generation of HAs during meat smoking processes and could be used as a reference to minimize the risks of cancer related to the consumption of smoked meat products.

Leuprolide is a synthetic nonapeptide drug (pyroGlu-His-Trp-Ser-Tyr-d-Leu-Leu-Arg-Pro-NHEt) that acts as a gonadotropin-releasing hormone agonist. The continuous administration of therapeutic doses of leuprolide inhibits gonadotropin secretion, which is used in androgen-deprivation therapy for the treatment of advanced prostate cancer, central precocious puberty, endometriosis, uterine fibroids, and other sex-hormone-related conditions. To improve the pharmacokinetic properties of peptide drugs, a fatty acid was conjugated with leuprolide for long-term action. In this study, we developed a simple ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for the simultaneous determination of leuprolide and leuprolide–oleic acid conjugate (LOC) levels. The developed method was validated in terms of linearity, precision, accuracy, recovery, matrix effect, and stability according to the US Food and Drug Administration guidelines, and the parameters were within acceptable limits. Subsequently, the pharmacokinetics of leuprolide and LOCs were evaluated. In vivo rat subcutaneous studies revealed that conjugation with fatty acids significantly altered the pharmacokinetics of leuprolide. After the subcutaneous administration of fatty-acid-conjugated leuprolide, the mean absorption time and half-life were prolonged. To the best of our knowledge, this is the first study showing the effects of fatty acid conjugates on the pharmacokinetics of leuprolide using a newly developed UPLC-MS/MS method for the simultaneous quantification of leuprolide and LOCs.

Funding Information: This work made use of the RawMatTERS Finland infrastructure (RAMI) facilities based at Aalto University. This work was a part of the Academy of Finland's Flagship Programme under Project No. 318890 and No. 318891 (Competence Center for Materials Bioeconomy, FinnCERES). Dr. Leena Pitkänen from Aalto University provided the assistance for the molecular weight analysis using Gel Permeation Chromatography. The NMR premises from the University of Helsinki is also appreciated. Publisher Copyright: © 2023 The Author(s) This study investigates the chemical structure of proanthocyanidin-rich crude extracts from willow bark and these materials were tested initially as adsorbents for artificial (waste)water treatment. The crude extracts were obtained through mild water extraction and the colorant fractions were further chromatographically fractionated to understand the chemical structure of the willow bark proanthocyanidins. The chemistry of crude extracts and purified fractions were investigated using nuclear magnetic resonance (NMR) and ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). Both NMR (liquid and solid-state) and UPLC-MS/MS suggest that the crude extracts constitute of interflavan linked flavan-3-ols, i.e. proanthocyanidins with both procyanidin (PC)-type and prodelphinidin (PD)-type subunits, with the PC/PD ratio of approximately 2.3–2.5. PD-type proanthocyanidins were detected from the purified colorant fractions only with UPLC-MS/MS. Both the UPLC-MS/MS and size exclusion chromatography suggest that the crude extracts have an average oligomerization degree of roughly 5–6 flavan-3-ol units. Adsorption experiments showed that the activated foams made of crude extracts were effective in removing both zinc and Bisphenol A (BPA) with removal efficiencies of roughly 80–90% and thus these willow bark-derived proanthocyanidins are promising in water treatment. The significance of this study suggests the upgrading use of crude extracts for water treatment could significantly improve the value of willow bark.

International audience; Isothiazolinones, a family of biocides, are used as preservatives for their fungicidal, bactericidal, and algacidal properties. These compounds can be found in a wide range of consumer and building products, such as paints, varnishes, shampoos, and liquid detergents. A robust analytical UPLC-MS/MS method to identify and quantify seven isothiazolinones (MIT, CMI, BIT, MBIT, BBIT, OIT, and DCOIT) in consumer products and their emissions was developed and validated according to ISO/IEC 17025. The limits of detection (LODs) ranged from 0.14 µg L-1 (BIT) to 0.54 µg L-1 (CMI). The method was applied for the quantification of the seven isothiazolinones in four types of consumer products (i.e., cosmetics, air fresheners, cleaning products, and building products) and the indoor emissions from a paint. Matrix effects were observed for the shampoo (63-74%), the shower gel (67-84%), and the ceramic glass cleaner (53-57%). All isothiazolinones indicated by the manufacturer (i.e., MIT, CMI, BIT, OIT, and DCOIT) were detected in the products and successfully quantified by the UPLC-MS/MS method.

(1) Background: Continuous monitoring of tacrolimus (TAC), mycophenolic acid (MPA), and creatinine (Cre) after renal transplantation is vitally important. In this study, we developed a new method based on volumetric absorptive microsampling (VAMS) combined with Ultra Performance Liquid Chromatography–Tandem Mass Spectrometry (UPLC-MS/MS) to simultaneously quantify three analytes including TAC, MPA, and Cre in whole blood. (2) Methods: The VAMS-based UPLC-MS/MS assay used a shared extraction and a single injection to simultaneously quantify the included TAC, MPA, and Cre. Development and validation were carried out following the Food and Drug Administration and European Medicines Agency guidelines for the validation of bioanalytical methods. Moreover, clinical validation for the three analytes was performed in both dried blood spot (DBS) and VAMS. Furthermore, a willingness survey was conducted using the system usability scale (SUS) for renal transplant recipients. (3) Results: The assay was successfully validated for all analytes. No interference, carryover, or matrix effects were observed, and extraction recoveries and process efficiencies were >90.00%. Analysis was unaffected by hematocrit (0.20~0.60, L/L) and anticoagulants (EDTA-2K). Dried VAMS samples were stable for 7 days at ambient temperature and stable for at least 1 month at −20 °C. During clinical validation, the measured TAC, corrected MPA, and Cre concentrations of VAMS samples met the analytical standards (95.00%, 88.57%, and 92.50%). When more stringent clinical acceptance criteria were set, the results obtained by VAMS (90.00%, 71.43%, and 85.00%) better than DBS (77.50%, 62.86%, and 70.00%). Compared with DBS, the survey found that renal transplant recipients are more inclined to use VAMS. (4) Conclusions: A robust extraction and UPLC-MS/MS analysis method in VAMS tips was developed and fully validated for the simultaneous quantification of TAC, MPA, and Cre concentrations. This method provides analytical support for the one-sample remote monitoring of both immunosuppressive drug concentrations and renal function in allo-renal recipients. Based on the good consistency between this method and the routine detection of venous blood samples and higher patient satisfaction than DBS, we believe that VAMS sampling can be a better alternative to venous whole-blood sampling.

Fagopyrum dibotrys, being native to southwest China, is widely distributed in Yunnan, Guizhou Provinces and Chongqing City. However, the quality of medicinal materials growing in different origins varies greatly, and cannot meet the market demand for high-quality F. dibotrys. In this study, 648 metabolites were identified, and phenolic compounds of F. dibotrys from different origins were clearly separated by principal component analysis (PCA). Our results suggested that the medicinal differences of F. dibotrys from different origins can be elucidated via the variations in the abundance of the phenolic and flavonoid compounds. We found that the epicatechin, total flavonoids and total tannin content in Yunnan Qujing (YQ) and Yunnan Kunming (YK) were higher than those in Chongqing Shizhu (CS), Chongqing Fuling (CF) and Guizhou Bijie (GB), suggesting that Yunnan Province can be considered as one of the areas that produce high-quality medicinal materials. Additionally, 1,6-di-O-galloyl-β-D-glucose, 2,3-di-O-galloyl-D-glucose and gallic acid could be used as ideal marker compounds for the quality control of F. dibotrys from different origins caused by metabolites, and the F. dibotrys planted in Yunnan Province is well worth exploiting.

Objective To develop an UPLC-MS/MS method for the accurate quantification of Estriol (E3), a new radioprotective agent, and observe the variation in pharmacokinetic characteristics of E3 after irradiation. As a hormone drug, the gender differences of E3 metabolism were also concerned here. Methods Various chromatographic and mass spectrometric conditions of E3 were optimized. The specificity, linearity, precision and accuracy of UPLC-MS/MS method were validated. Twenty SD rats, half male and half female, were administered with intramuscular (im) injection of 2.7 ​mg/kg E3, and then divided randomly into two groups, sham-irradiated group (Con) and irradiated group (IR). IR group was irradiated to 7 ​Gy of γ-rays. Blood samples were collected at different times post-irradiation and E3 concentrations were detected. The changes of concentration-time variation and pharmacokinetic parameters for E3 after irradiation were investigated, together with metabolic differences between male and female rats. Results The range of the calibration curve of UPLC-MS/MS for E3 was 1.00–200.0 ​ng/mL. Con group reached maximum concentration (Cmax) (77.57 ​± ​18.71) ng/mL at (0.68 ​± ​0.29) h (Tmax) after im injection. The drug concentration-time profiles and pharmacokinetic parameters of E3 were consistent before and after irradiation. The areas under time curve (AUC0-t) of E3 were (353 ​± ​74) h·ng/mL for Con group, and (299 ​± ​74) h·ng/mL for IR group (P ​> ​0.05). There were also no statistical differences in pharmacokinetic parameters between female and male rats. The elimination half-lifes (T1/2) of males and females were (3.02 ​± ​0.68) h and (3.01 ​± ​0.42) h in Con group, and (3.64 ​± ​0.51) h and (3.38 ​± ​0.60) h in IR group, respectively (P ​> ​0.05). Conclusion A rapid and sensitive UPLC-MS/MS method for determination of E3 was established. The pharmacokinetic characteristics of E3 in rat were not affected by 7 ​Gy irradiation and gender differences. This study provided a theoretical basis for the development and application of new radiation injury treatment drug.

Suili Yang,1,* Xiaoshan Zhang,2,3,* Yuzhen Wang,2,3 Congcong Wen,4 Chenxiang Wang,3 Ziye Zhou,5 Guanyang Lin3 1Department of Neurology, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, Zhejiang, People’s Republic of China; 2College of Pharmacy, Wenzhou Medical University, Wenzhou, Zhejiang, People’s Republic of China; 3Department of Pharmacy, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, Zhejiang, People’s Republic of China; 4Laboratory Animal Center, Wenzhou Medical University, Wenzhou, Zhejiang, People’s Republic of China; 5Clinical Research Center, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, Zhejiang, People’s Republic of China*These authors contributed equally to this workCorrespondence: Guanyang Lin; Ziye ZhouDepartment of Pharmacy, The First Affiliated Hospital of Wenzhou Medical University, Nanbaixiang Street, Ouhai District, Wenzhou City, 325000, People’s Republic of ChinaTel +86-577-55579706Email 13867702133@163.com; redd88@163.comBackground and Aim: Dasatinib is approved for the treatment of leukaemia worldwide. Triazole agents such as posaconazole may be used for the control of secondary fungal infection with leukaemia. This work aimed to develop a bioanalytical method to study the potential interaction between dasatinib and posaconazole.Methods: An ultrahigh-performance liquid chromatography-tandem mass spectrometry method was established to measure the plasma concentrations of dasatinib and posaconazole in rats simultaneously. Simple protein precipitation with acetonitrile was applied to extract dasatinib and posaconazole in samples. The chromatographic separation of analytes was conducted on an UPLC BEH C18 column using a mobile phase consisting of 0.1% aqueous formic acid and acetonitrile. Dasatinib and posaconazole were monitored in positive ion mode with the following mass transition pairs: m/z 488.2→ 401.1 for dasatinib and m/z 701.3→ 683.4 for posaconazole. The method was successfully applied for pharmacokinetic interaction between dasatinib and posaconazole.Results: The established method expressed good linearity in 1– 1000 ng/mL of dasatinib and 5– 5000 ng/mL of posaconazole, with limit of detection was 1 ng/mL and 5 ng/mL, respectively. Methodology validations, including accuracy, precision, matrix effect, recovery, and stability, met the US Food and Drug Administration (FDA) acceptance criteria for bioanalytical method validation. Dasatinib strongly inhibited the clearance of posaconazole in vivo, while posaconazole expressed no significant effect on the pharmacokinetics of dasatinib.Conclusion: Dasatinib alters the pharmacokinetics of posaconazole. Attention should be paid to the unexpected risk of adverse clinical outcomes when posaconazole is co-administered with dasatinib.Keywords: dasatinib, posaconazole, UPLC-MS/MS, interaction, pharmacokinetics

Chen-jian Zhou,1 Hui-jun Wang,2 Chun-yan Zhou,2 Chao-fan Li,2 Ming-Jia Zhu,2 Xiang-jun Qiu2 1Department of Pharmacy, Wenzhou Central Hospital, Wenzhou, 325027, People’s Republic of China; 2School of Basic Medical Sciences, Henan University of Science and Technology, Luoyang, 471023, People’s Republic of ChinaCorrespondence: Xiang-jun QiuSchool of Basic Medicine, Henan University of Science and Technology, 263 Kaiyuan Avenue, Luoyang, 471023, People’s Republic of ChinaTel +8613698882699Fax +86379-64830346Email lyxiangjun@126.comBackground: A method for the determination of selinexor by UPLC-MS/MS was established to study the effect of posaconazole on the pharmacokinetics of selinexor in rats.Methods: The experiment rats were divided into group A (0.5% CMC-Na) and group B (posaconazole, 20 mg/kg), 6 rats in each group. 30 minutes after administration of 0.5% CMC-Na or posaconazole, all the rats were given selinexor (8 mg/kg), and plasma samples were collected. The plasma samples underwent acetonitrile protein precipitation, and were separated by UPLC on an Acquity UPLC BEH C18 column with gradient elution. Acetonitrile and 0.1% formic acid were used as the mobile phases. The analyte detection was used a Xevo TQ-S triple quadrupole tandem mass spectrometer and multiple reaction monitoring (MRM) for analyte monitoring. We use acetonitrile for protein precipitation.Results: Selinexor had good linearity (1.0– 1000 ng/mL, r2= 0.996 2), and the accuracy and precision, recovery rate and matrix effects(ME) were also met the FDA approval guidelines. Compared with group A, the Cmax, AUC(0−t) and AUC(0−∞) of selinexor in group B increased by 60.33%, 48.28% and 48.27%, and Tmax increased by 53.92%, CLz/F reduced by 32.08%.Conclusion: This bioanalysis method had been applied to the study of drug interactions in rats. It was found that posaconazole significantly increased the concentration of selinexor in rats. Therefore, when selinexor and posaconazole are combined, we should pay attention to the possible drug–drug interactions to reduce adverse reactions.Keywords: UPLC-MS/MS, selinexor, posaconazole, pharmacokinetics, DDIs, rats

Fenebrutinib (GDC-0853; FNB) is an oral small molecule that was developed by Roche Pharmaceuticals to slow multiple sclerosis progression. FNB is a reversible bruton tyrosine kinase (BTK) inhibitor, which showed the maximum potency of BTK inhibitors in phase III clinical trials for multiple sclerosis. In the current study, a fast, specific, and sensitive UPLC-MS/MS method for FNB quantification in human liver microsomes (HLMs) was established with application to the evaluation of metabolic stability. The UPLC-MS/MS methodology was verified using the stated USFDA validation guidelines for bioanalytical methodologies that involve selectivity, linearity, accuracy and precision, carryover and extraction recovery, stability, and matrix effect. The FNB calibration curve displayed a linearity in the range from 1 ng/mL to 3000 ng/mL (y = 1.731x + 2.013; R2: 0.9954; RSD < 4.37%) in the HLMs matrix. The limit of quantification was 0.88 ng/mL, which verified the UPLC-MS/MS analytical method sensitivity. The intraday and interday precision and accuracy results of the developed UPLC-MS/MS method were −3.99–14.0% and 0.52–3.83%, respectively. FNB and savolitinib (SVB) (internal standard) were chromatographically separated utilizing an isocratic mobile phase system with a ZORBAX Eclipse plus-C18 (50 mm, 2.1 mm, and 1.8 μm) column. The metabolic stability parameters for FNB, involving high intrinsic clearance (58.21 mL/min/kg) and a short in vitro half-life (13.93 min), revealed the high extraction ratio of FNB. Reviewing the literature revealed that the current UPLC-MS/MS method is the first analytical method for FNB quantification in the HLMs matrix with application to the assessment of FNB metabolic stability.

TMAO is a new risk biomarker for cardiovascular disease. With trimethylammonium as its main chemical skeleton, TMAO is structurally similar to many endogenous metabolites, such as acetylcholine, carnitine, phosphorylcholine, etc. The mechanism of TMAO on the pathological process of CVD is still unclear. In this study, the quantitative analysis of plasma TMAO is conducted, and the contribution of Cathepsin B and NLRP3 inflammasome during the process of TMAO-induced endothelial injury was proposed and investigated at animal and cellular levels. Immunofluorescence assay was applied to represent the protein expression of Cathepsin B and NLRP3 inflammasome located at endothelial cells. The results showed that TMAO could disrupt endothelial cells permeability to induce endothelial injury, meanwhile, TMAO could increase NLRP3 inflammasome activation and promote the activity and expression of Cathepsin B in vitro and in vivo, whereas inhibition of NLRP3 inflammasome activation by MCC950 could protect the endothelial cells from TMAO associated endothelial injury via Cathepsin B. The study reveals that TMAO can cause endothelial injury via Cathepsin B-dependent NLRP3 inflammasome, and inhibition of Cathepsin B and NLRP3 inflammasome can reduce the TMAO-induced damage. The results provide new insight into the role of TMAO in CVD, which can be a potential therapeutic target for disease treatment and drug design.

Alkylphenol ethoxylates (APEOs) represent a non-ionic surfactant widely used as adjuvants in pesticide formulation, which is considered to cause an endocrine-disrupting effect. In the current study, we established a detection method for the APEOs residue in tea based on solid-phase extraction (SPE) for the simultaneous analysis of nonylphenol ethoxylates (NPEOs) and octylphenol ethoxylates (OPEOs) by UPLC–MS/MS. In the spiked concentrations from 0.024 to 125.38 μg/kg for 36 monomers of APEOs (nEO = 3–20), the recoveries of APEOs range from 70.3–110.7% with RSD ≤ 16.9%, except for OPEO20 (61.8%) and NPEO20 (62.9%). The LOQs of OPEOs and NPEOs are 0.024–6.27 and 0.16–5.01 μg/kg, respectively. OPEOs and NPEOs are detected in 50 marketed tea samples with a total concentration of 0.057–12.94 and 0.30–215.89 µg/kg, respectively. The detection rate and the range of the monomers of NPEOs are generally higher than those of OPEOs. The current study provides a theoretical basis for the rational use of APEOs as adjuvants in commercial pesticide production.

Gabapentin (GAB) is a cyclohexane acetic acid, structurally related to the neurotransmitter gamma-aminobutyric acid (GABA), and considered the principal inhibitory neurotransmitter in the central nervous system (CNS) of mammals. An ultra-performance liquid chromatography–tandem mass spectrophotometry (UPLC-MS/MS) method for assessing pregabalin (PRE) in human plasma, was developed and validated, via PRE usage as an internal standard. The plasma underwent protein precipitation using methanol, prior to analysis. Chromatographic separation was completed using a mobile phase of methanol: 0.1% formic acid solution, (65:35, v/v), at a flow rate of 0.2 mL/min, with an isocratic approach, on an Agilent Eclipse plus column (50 × 2.1 mm and 1.8 μm), in 1.6 min of running time. An Agilent triple quadrupole was used for mass analysis, to detect the ion transitions for GAB and PER, respectively, at m/z of 172.1 → 154.1 and 160.10 → 142.10. The calibration curve, over the linear range of 0.050–10.0 μg/mL, showed a high correlation coefficient, r = 0.9993. The limits of detection and quantitation were 13.37 ng/mL and 40.52 ng/mL, respectively, based on the standard deviation and slope equation. The results for intra- and inter-day measurement accuracy and precision were in acceptable ranges. The method was extended into the assessment of oral administrations of GAB at different doses, of one 600 mg/tablet and two capsules (each one of them has 300 mg of GAB), to volunteers who were used in pharmacokinetics and bioequivalent studies. The AGREE assessment tool was used to visualize the proposed method’s greenness degree, which revealed a high AGREE rating score, supporting the accepted method’s greenness profile.

Bisphenol compounds (BPs) are of concern worldwide as potential environmental contaminants, and these chemicals may migrate into animal-derived foodstuffs from animal feeds. In this study, 12 BPs were analyzed in 7 varieties of animal feed groups totaling 203 samples collected from various provinces in China. 6 BPs were found in these feed samples, with a total detection frequency of 75.5%. The total concentration of all BPs (Σ6BPs) in each type of feed ranged from below the limit of detection to 75.3 ng g−1, with an overall mean value of 4.52 ng g−1. The frequency of BPs contamination and the Σ6BPs values were found to be higher in animal-based feeds (Pfrequency > 0.05, Pconcentration < 0.01) than in plant-based feeds. BPA and BPS were the predominant BPs in these feed samples, accounting for (on average) 63.9% and 24.6% of the total BPs, respectively. Overall, the mean concentrations of BPA in these animal feeds were much lower than the legal limits set by China and European Union, implying a low direct risk to both animal health and animal-derived food safety. However, the evident risk of the contamination of various animal-derived food items with BPs passed through the feed chain merits further investigation.

Accurate DNA quantitation is a prerequisite in many biomedical and pharmaceutical studies. Here we established a new DNA quantitation method by nuclease P1 digestion and UPLC-MS/MS analysis. DNA fragments can be efficiently hydrolyzed to single deoxyribonucleotides by nuclease P1 in a short time. The decent stabilities of all the four deoxyribonucleotides were confirmed under different conditions. Deoxyadenosine monophosphate (dAMP) was selected as the surrogate for DNA quantitation because dAMP showed the highest sensitivity among the four deoxyribonucleotides in the UPLC-MS/MS analysis. The linear range in DNA quantitation by this method is 1.2–5000 ng/mL. In the validation, the inter-day and intra-day accuracies were within 90%–110%, and the inter-day and intra-day precision were acceptable (RSD, Graphical abstract Image 1, Highlights • A novel method to evaluate DNA binding efficiency of the DNA alkylator PBD in tumors of mouse models is reported. • The DNA isolation, DNA digestion and the LC/MS quantitation of both DNA and PBD are involved. • The method with an enhanced sensitivity allows for the accurate quantitation of isolated DNA from various tissues.

Foretinib, an oral multikinase inhibitor, is known to have anti-tumor effects against cancers. The doses and the levels of foretinib vary based on the type of cancer to be treated. An accurate and precise method is required to determine the level of foretinib and its pharmacokinetics. Here, we developed such a method, which was validated based on the guidelines of the FDA and EMA. Foretinib and ibrutinib (the internal standard (IS)) were extracted using tert-butyl methyl ether. Foretinib and IS were eluted in approximately 1.2 min. Thus, a linear, fast, accurate, and precise method was developed. The calibration curve was linear (r2 ˃ 0.997) in the range of 0.5–400.0 ng/mL and the lowest limit of quantitation was 0.5 ng/mL. The average recovery, accuracy, and precision were 87.9%, 88.7%, and ≤7.8%, respectively. The analyte was deemed stable using various stability tests. The validated assay was then fruitfully applied to a pharmacokinetics study in rats, which revealed that foretinib was absorbed and the maximum concentration achieved at 4.0 h after the administration of a single dose of foretinib. Keywords: UPLC–MS/MS, Foretinib, Pharmacokinetic, Rats, Ibrutinib

Shuang-long Li, Yong-liang Zhu, Chun-yang Zhu, Shao-bin Li, Zi-heng Li, Xiang-jun Qiu Medical College of Henan University of Science and Technology, Luoyang 471023, People’s Republic of ChinaCorrespondence: Xiang-jun QiuMedical College of Henan University of Science and Technology 471023, People’s Republic of ChinaTel +86 13698882699 Fax + 86 379-64830346 Email lyxiangjun@126.comAbstract: A method for the simultaneous determination of parecoxib and its metabolite valdecoxib in beagle plasma by UPLC-MS/MS was developed and validated. After the plasma was extracted by acetonitrile precipitation, the analytes were separated on an Acquity UPLC BEH C18 column (2.1 mm × 50 mm, 1.7 μm) using acetonitrile-formic acid as the mobile phase in gradient mode. The analytes were monitored by multiple reaction monitoring (MRM) in electrospray negative ion mode. The mass transfer pairs were m/z 368.97→ 119.01 for parecoxib, m/z 312.89→ 118.02 for valdecoxib, and m/z 379.98→ 316.02 for celecoxib (internal standard, IS). The correlation coefficients of parecoxib and valdecoxib ranged from 5to 4000 ng/mL were greater than 0.9998. The recovery of parecoxib and valdecoxib was greater than 82.54%. The inter- and intra-day precision RSD values were 1.36∼ 3.65% and 2.28∼ 5.91%, respectively. The accuracy ofRE values were − 1.38%∼ 1.96%. Finally, the matrix effect (ME) and stability were also within acceptable criteria. This method had been successfully applied to the pharmacokinetics of parecoxib and valdecoxib in beagle plasma after injection of parecoxib (1.33 mg/kg, intramuscular injection).Keywords: parecoxib, valdecoxib, UPLC-MS/MS, pharmacokinetics

As set in the maximum residue limit regulations of the European Commission, this study aimed to obtain the residual parameters in milk with optimized UPLC-MS/MS conditions and to determine the conclusive drug withdrawal period to ensure food safety. In this research, an ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed to study cefquinome sulfate’s residue elimination in milk and to calculate cefquinome’s withdrawal period. Twelve healthy cows free of endometritis were selected for the experiment. Before using the drug, the vaginal orifice and perineum of each cow was disinfected. One dose of intrauterine perfusion was used for each cow, followed by an additional dose after 72 h. Before administration and 12 h, 18 h, 24 h, 36 h, 42 h, 48 h, 60 h, 66 h, 72 h, 84 h, 90 h, and 96 h after the last dose, milk (10 mL) was gathered from each cow’s teat and pooled. For the measurement of cefquinome concentrations in milk, UPLC-MS/MS was performed. A calibration curve was generated using linear regression as follows: Y = 250.86X − 102.29, with a correlation coefficient of 0.9996; the limits of detection and the limits of quantitation were 0.1 μg·kg−1 and 0.2 μg·kg−1, respectively. The average recovery of cefquinome was 88.60 ± 16.33% at 0.2 μg·kg−1, 100.95 ± 2.54% at 10 μg·kg−1, and 97.29 ± 1.77% at 50 μg·kg−1. For 5 consecutive days at the three spiking levels, the intra and inter-day relative standard deviations (RSD) were 1.28%–13.73% and 1.81%–18.44%, respectively; the residual amount of cefquinome was less than the maximum residue limit of 20 μg·kg−1, 36 h after administration; and the residual amount was less than the limit of detection (0.1 μg·kg−1) 48 h after administration. The withdrawal time of cefquinome in cow’s milk was 39.8 h, as calculated using WTM1.4 software. In terms of clinical practical use, the withdrawal period of milk was temporarily set at 48 h after the administration of the cefquinome sulfate uterus injection to cows, in accordance with the recommended dose and course.

Panax ginseng (P. ginseng), the dried root and rhizome of P. ginseng C. A. Meyer, is widely used in many fields as dietary supplements and medicine. To characterize the chemical constituents in P. ginseng cultivated in different growth environments, a UPLC-TOF-MS method was established for qualitative analysis. Four hundred and eight ginsenosides, including 81 new compounds, were characterized in P. ginseng from different regions. Among the detected compounds, 361 ginsenosides were recognized in P. ginseng cultivated in the region of Monsoon Climate of Medium Latitudes, possessing the largest amount of ginsenosides in all samples. Furthermore, 41 ginsenosides in 12 batches of P. ginsengs were quantified with a UPLC-MRM-MS method, and P. ginsengs from different regions were distinguished via chemometric analysis. This study showed that the different environments have a greater influence on P. ginseng, which laid a foundation for further quality control of the herb.

Robust and sensitive ultrahigh-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) combined with the quick, easy, cheap, effective, rugged, and safe (QuEChERS) method was applied for the detection of seven Alternaria toxins (ATs) in tuberous crops. The influence of tuber conditions (fresh, germinated, and moldy) during storage on the concentration of the seven ATs is also investigated. ATs were extracted with acetonitrile under acidic conditions and purified with a C18 adsorbent. ATs were scanned with electrospray ionization (positive/negative ion) dynamic switching and detected in MRM mode. Calibration curve analysis results reveal good linear relationships in all toxin concentration ranges (R2 > 0.99). The limit of detection and limit of quantification were 0.25–0.70 and 0.83–2.31 μg/kg, respectively. The average recoveries of the seven ATs were 83.2–104% with intra-/inter-day precision at 3.52–6.55% and 4.02–7.26%, respectively. The developed method provided adequate selectivity, sensitivity, and precision in detecting the seven ATs at trace levels, and dispensed with standard addition or matrix-matched calibration to compensate for matrix effects. ATs in the fresh, germinated, and moldy samples of tuberous crops in storage (taro, potato, sweet potato, yam, cassava) were analyzed with this method, and the concentrations were 2.01–14.51 μg/kg and significantly increased with storage duration. ALS was detected in most samples, whereas no quantities of ALT and ATX-I were detected. AME was often detected in combination with AOH in sweet potatoes. TeA and Ten were mostly detected in taro, potato, and yam. The established method could be used for the simultaneous detection and quantification of multicomponent toxins in elaborate matrices.

Primula veris subsp. veris L. is a perennial herbaceous and medicinal plant species the roots and flowers of which are a source of valuable pharmaceutical raw materials. The plant tissues are used to produce expectorant and diuretic drugs due to their high content of triterpene saponins and phenolic glycosides. Underground roots of P. veris can be obtained only through a destructive process during the plant’s harvesting. In the present study, an in vitro adventitious root production protocol was developed as an alternative way of production, focused on four species-specific secondary metabolites. Root explants were cultured in Murashing & Skoog liquid medium supplemented with 5.4 μM α-naphthaleneacetic acid, 0.5 μM kinetin, L-proline 100 mg/L, and 30 g/L sucrose, in the dark and under agitation. The effect of temperature (10, 15 and 22 °C) on biomass production was investigated. The content of two flavonoid compounds (primeverin and primulaverin), and two main triterpene saponins (primulic acid I and II) were determined after 60 days of culture and compared with 1.5-year-old soil-grown plants. The accumulated content (mg/g DW) of bioactive compounds of in vitro adventitious roots cultured under 22 °C was significantly higher than the other two temperatures of the study, being 9.71 mg/g DW in primulaverin, 0.09 mg/g DW in primeverin, 6.09 mg/g DW in primulic acid I, and 0.51 mg/g DW in primulic acid II. Compared to the soil-grown roots (10.23 mg/g DW primulaverin, 0.28 mg/g DW primeverin, 17.01 mg/g DW primulic acid I, 0.09 mg/g DW primulic acid II), the in vitro grown roots at 22 °C exhibited a 5.67-fold higher content in primulic acid II. However, primulic acid I and primeverin content were approximately three-fold higher in soil-grown roots, while primulaverin content were at similar levels for both in vitro at 22 °C and soil-grown roots. From our results, tissue culture of P. veris subsp. veris could serve not only for propagation but also for production of species-specific secondary metabolites such as primulic acid II through adventitious root cultures. This would therefore limit the uncontrolled collection of this plant from its natural environment and provide natural products free from pesticides in a sustainable way.

Parabens are endocrine-disrupting chemicals (EDCs) that have estrogen-like activities and may cause male reproductive disorders. Here, we developed a method for the simultaneous determination of four parabens (MeP, EtP, n-PrP, n-BuP) and two metabolites (4-HB and 3,4-DHB) in human seminal plasma by UPLC-MS/MS. The method was used to analyze 144 seminal plasma samples from Chinese males. MeP, EtP, n-PrP, and 4-HB were the dominant compounds. MeP, EtP, and n-PrP were significantly correlated to each other. In addition, 4-HB was significantly correlated to MeP, EtP, n-PrP, and 3,4-DHB, respectively. The results provide direct evidence that parabens and their metabolites are widely distributed in the male reproductive system. The study presents the paraben metabolites levels in human seminal plasma for the first time.

Here, a QuEChERS (quick, easy, cheap, effective, rugged, and safe) pretreatment method was combined with UPLC-MS/MS to facilitate the rapid and reliable simultaneous detection of five calcium channel blockers (CCBs) in human plasma. For this approach, samples were treated with 1 mL of acetonitrile, 350 mg of magnesium sulfate, and 70 mg of PSA adsorbent prior to centrifugation. Supernatants then underwent gradient elution for 8 min with an Agilent C18 column using an acetonitrile-water solution supplemented with 5 mmol⋅L−1 of ammonium acetate. This technique exhibited a good linear response in the 1–800 ng⋅mL−1 range for the analyzed drugs, with an R2≥ 0.9921, an accuracy of 87.54–113.05%, a matrix effect (ME) of 91.21–116.39%, a precision of 0.19–11.64%, and stability of no more than 10.05%. This time-saving QuEChERS reagent-based pretreatment technique thus allowed for the simultaneous and accurate detection of five CCBs in human plasma samples, providing a promising new basis for therapeutic drug monitoring in patients with hypertension.

Chlorfenapyr, as a highly effective and low-toxicity insect growth regulation inhibitor, has been used to control cross-cruciferous vegetable pests. However, the pesticide residue caused by its application threatens human health. In this paper, the residue digestion and final residue of chlorfenapyr in radish were studied in a field experiment. The results of the dynamic digestion test showed that the half-life of chlorfenapyr in radish leaves ranged from 6.0 to 6.4 days, and the digestion rate was fast. The median residual values of chlorfenapyr in radish and radish leaves at 14 days after treatment were 0.12 and 3.92 mg/kg, respectively. The results of the dietary intake risk assessment showed that the national estimated daily intake (NEDI) of chlorfenapyr in various populations in China were 0.373 and 5.66 µg/(kg bw·d), respectively. The risk entropy (RQ) was 0.012 and 0.147, respectively, indicating that the chronic dietary intake risk of chlorfenapyr in radish was low. The results of this study provided data support and a theoretical basis for guiding the scientific use of chlorfenapyr in radish production and evaluating the dietary risk of chlorfenapyr in vegetables.

Diatoms are a successful group of microalgae at the base of the marine food web. For hundreds of millions of years, they have shared common habitats with bacteria, which favored the onset of interactions at different levels, potentially driving the synthesis of biologically active molecules. To unveil their presence, we sequenced the genomes of bacteria associated with the centric diatom Thalassiosira rotula from the Gulf of Naples. Annotation of the metagenome and its analysis allowed the reconstruction of three bacterial genomes that belong to currently undescribed species. Their investigation showed the existence of novel gene clusters coding for new polyketide molecules, antibiotics, antibiotic-resistance genes and an ectoine production pathway. Real-time PCR was used to investigate the association of these bacteria with three different diatom clones and revealed their preference for T. rotula FE80 and Skeletonema marinoi FE7, but not S. marinoi FE60 from the North Adriatic Sea. Additionally, we demonstrate that although all three bacteria could be detected in the culture supernatant (free-living), their number is up to 45 times higher in the cell associated fraction, suggesting a close association between these bacteria and their host. We demonstrate that axenic cultures of T. rotula are unable to grow in medium with low salinity (

Substantial efforts have been made in incorporating tannin-rich forages into grassland-based livestock production systems. However, the structural and functional diversity of tannins in different species limits their potential use at the field scale. We conducted a greenhouse experiment with 17 cultivars from 8 forage species and their cultivars. Ultraperformance liquid chromatography tandem mass spectrometry was used to analyze their polyphenolic profile and proanthocyanidin (PA) structural features in leaves. Our results highlight large inter- and intraspecies variability of plants in terms of polyphenol and tannin concentrations in the leaves. A concomitant and significant variation was also registered in the structural features of PA-rich forages such as the mean degree of polymerization and prodelphinidin percentage. The concentration of PA also varied within plant organs; the highest concentration was in flowers, but leaves had the highest contribution to harvestable PA biomass. Our research highlights that identifying these variations helps in identifying the representativeness of bioactivity and provides the basis for targeted breeding programs.

Lonicera maackii (Caprifoliaceae) is a large, upright shrub with fruits that contain many bioactive compounds. Flavonoids are common active substances in L. maackii. However, there is a dearth of information about the accumulation of these flavonoids and their possible medicinal value. We used targeted metabolomics analysis based on ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) to analyze five developmental stages of L. maackii fruit. A total of 438 metabolites were identified in the five developmental stages, including 81 flavonoids and derivatives. The 81 flavonoids included 25 flavones and derivatives, 35 flavonols and derivatives, two isoflavones, three cyanidins and derivatives, eight procyanidins, and eight flavanones. In addition, we outlined the putative flavonoid biosynthesis pathway and screened their upstream metabolites. More importantly, we analyzed the accumulation patterns of several typical flavones and flavonols. The results reported here improved our understanding of the dynamic changes in flavonoids during fruit development and contributed to making full use of the medicinal value of L. maackii fruit.

Ginger (Zingiber officinale Roscoe) is known for its unique pungent taste and useability in traditional Chinese medicine. The main compounds in ginger rhizome can be classified as gingerols, diarylheptanoids, and volatile oils. The composition and concentrations of the bioactive compounds in ginger rhizome might vary according to the age of the rhizome. In this regard, the knowledge on the transcriptomic signatures and accumulation of metabolites in young (Y), mature (M), and old (O) ginger rhizomes is scarce. This study used HiSeq Illumina Sequencing and UPLC-MS/MS analyses to delineate how the expression of key genes changes in Y, M, and O ginger rhizome tissues and how it affects the accumulation of metabolites in key pathways. The transcriptome sequencing identified 238,157 genes of which 13,976, 11,243, and 24,498 were differentially expressed (DEGs) in Y vs. M, M vs. O, and Y vs. O, respectively. These DEGs were significantly enriched in stilbenoid, diarylheptanoid, and gingerol biosynthesis, phenylpropanoid biosynthesis, plant-hormone signal transduction, starch and sucrose metabolism, linoleic acid metabolism, and α-linoleic acid metabolism pathways. The metabolome profiling identified 661 metabolites of which 311, 386, and 296 metabolites were differentially accumulated in Y vs. M, Y vs. O, and M vs. O, respectively. These metabolites were also enriched in the pathways mentioned above. The DEGs and DAMs enrichment showed that the gingerol content is higher in Y rhizome, whereas the Y, M, and O tissues differ in linoleic and α-linoleic acid accumulation. Similarly, the starch and sucrose metabolism pathway is variably regulated in Y, M, and O rhizome tissues. Our results showed that ginger rhizome growth slows down (Y > M > O) probably due to changes in phytohormone signaling. Young ginger rhizome is the most transcriptionally and metabolically active tissue as compared to M and O. The transitioning from Y to M and O affects the gingerol, sugars, linoleic acid, and α-linoleic acid concentrations and related gene expressions.

Tamoxifen, a widely prescribed medication in premenopausal women diagnosed with hormone-dependent breast cancer, is potentially co-prescribed with Hedyotis diffusa (H. diffusa), particularly in Taiwan. However, no related report has investigated the drug-herb interaction of H. diffusa on the pharmacokinetics of tamoxifen and its metabolites. In the present study, male Sprague-Dawley rats were administered different doses of H. diffusa extract for 5 consecutive days prior to the administration of tamoxifen (10 mg/kg). A validated ultra-liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) system was developed to monitor tamoxifen, 4-hydroxytamoxifen, N-desmethyltamoxifen, and endoxifen in rat plasma. Pharmacokinetic results demonstrated that the area under curves (AUCs) of tamoxifen and the relative bioavailability (%) of tamoxifen were dose-dependently decreased (31–68%) by pre-treatment with H. diffusa extract (3 g/kg and 6 g/kg). In addition, the conversion ratio of 4-hydroxytamoxifen was downregulated (0.5-fold change) and the N-desmethyltamoxifen conversion ratio was upregulated (2-fold change) by high-dose H. diffusa extract. As a result, the relative bioavailability and biotransformation changes affect the clinical efficacy of tamoxifen treatment. These preclinical findings reveal a hitherto unreported interaction between tamoxifen and H. diffusa extract that has implications for their therapeutic efficacy in treating breast cancer.

Highlights • Lactose in human milk affects B-vitamin analysis by fouling the ion source. • Typical extraction methods are inefficient for lactose removal. • Chromatographic separation from analytes enables lactose mechanical removal., Our previously reported, first validated, UPLC-MS/MS-based simultaneous analysis of five human milk B-vitamins revealed severe matrix effects. High levels of endogenous lactose fouled the electrospray ionization source affecting the analysis. We evaluated solid-phase extraction (SPE), liquid–solid extraction (LSE), protein precipitation (PPT), and liquid chromatography effluent diversion for lactose-removal. SPE failed to separate lactose from vitamins; LSE using 2-propanol reduced lactose and vitamin recoveries. PPT-solvent, milk volume, and reconstitution solvent influenced flavin adenine dinucleotide, pyridoxal and nicotinamide recoveries. Using an optimized LC-gradient enabled chromatographic separation of lactose from vitamins and its removal using a post-column switch-valve. Only 40 µL milk was subjected to methanol-PPT and non-polar matrix removal by methyl tert-butyl ether. B-vitamin recoveries were established (81.9–118.6%; CV ≤ 11.9%; precision: 4.9–13.7%) with greatly reduced matrix effects, and improved process efficiency, and recovery.

In this study, an ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed for the residue depletion of imidocarb (IMD) in bovine tissues, and the drug withdrawal time of IMD was determined. Twenty-five clinically healthy cattle (body weight 300 kg ± 15 kg) were randomly divided into five groups of five cattle each. The cattle were treated subcutaneously injecting a single dose of a generic IMD formulation, at the recommended dosage of 3.0 mg/kg. The five groups of cattle were slaughtered respectively at 96, 160, 198, 213, and 228 days after IMD administration. Samples from the liver, kidney, muscle, fat, and injection site were collected from each animal. After subtilis proteinase was used to digest the tissue, the content of IMD in the samples was analyzed by UPLC-MS/MS method. In conclusion, the method validation results showed that the method meets the criteria, and the longest withdrawal time of 224 days for the liver can be selected as the conclusive withdrawal time to guarantee consumer safety.

Biogenic amines (BAs) are a group of substances that are formed from amino acids by decarboxylation or amination and transamination of aldehydes and ketones. They may have either an aliphatic, aromatic, or heterocyclic structure. Their quantity determines their effects and optimum amounts are essential for physiological functions, but excess BAs causes various toxic effects throughout the human body. In our study, to rapidly determine 14 BAs (histamine, tyramine, dopamine, tryptamine, serotonin, putrescine, spermine, spermidine, octopamine, benzylamine, 1-Phenylethanamine, cadaverine, 2-Phenethylamine, and agmatine) in real fish samples, an ultra-performance liquid chromatography–tandem mass spectrometry method was established. The fish sample was extracted by acetonitrile with 0.1% formic acid and stable biogenic amine derivatives could be obtained by benzoyl chloride derivatization with a shorter reaction time. The method showed good linearity with a linear range of 3–4 orders of magnitude and regression coefficients ranging from 0.9961 to 0.9999. The calculated LODs ranged from 0.1 to 20 nM and the LOQs ranged from 0.3 to 60 nM. Satisfactory recovery was obtained from 84.6% to 119.3%. The proposed method was employed to determine the concentration levels of biogenic amine derivatives in different fish. The results indicated that this method was suitable for the analysis of biogenic amines.

Tetrodotoxin (TTX) is a kind of low-molecular-weight non-protein neurotoxin. It is one of the most potent neurotoxins found in nature, and it is found in puffer fish and various marine biota. The low sensitivity of previous analytical methods limited their application in puffer fish organ samples. This study established a method for the accurate and fast determination of TTX by reversed ultra-performance liquid chromatography coupled with proton-enhanced electron spray ionization–tandem mass spectrometry. The method yields good peak shapes, high sensitivity and low coeluted interferences. The method was successfully applied to determine TTX in puffer fish tissue samples of about 0.2 g.

Recently, neoadjuvant treatment has turned out to be a feasible alternative for individuals suffering from locally advanced breast cancer. The neoadjuvant therapy is a type of chemotherapy that is given either before or after surgeries to diminish a tumor and minimize the likelihood of recurrence. This article demonstrates the development of a unique bioanalytical validated sensitive method by means of an ultra high performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) approach for the concurrent estimation of neoadjuvant treatments including 5-Fluorouracil, Doxorubicin, and Capecitabine in rat plasma. Samples were prepared using the fine minor QuEChERS process and analyzed using a Shimadzu-C18 column via an isocratic separation. Acetonitrile:water in the ratio of (30:70) (both containing 0.1 percent formic acid v/v) was the mobile phase employed at a flow rate of 0.20 mL/min. At concentrations of 50.00–500.00 ng/mL for 5-Fluorouracil, 25.00–500.00 ng/mL for Doxorubicin, and 5.00–100.00 ng/mL for Capecitabine, the procedure was shown to be linear. The limit of detection (LOD) was assessed in ng/mL and varied from 1.33 to 13.50. Relative standard deviations for precision were below 2.47 percent over the whole concentration range. For all analytes, the average recovery rate varied from 73.79 to 116.98 percent. A preliminary pharmacokinetic study was successfully performed in real rats to evaluate the procedure efficiency.

Antibiotic distribution and analysis within liquid and solid fractions of manure are highly variable due to each compound’s respective physiochemical properties. This study developed and evaluated a uniform method extracting 10 antibiotics from 4 antibiotic classes (tetracycline, sulfonamides, macrolides, and β-lactam) from unprocessed manure, solid–liquid separated manure, and composted solids. Through systematic manipulation of previously published liquid chromatography tandem mass spectrometry methods; this study developed an extraction protocol with optimized recovery efficiencies for varied manure substrates. The method includes a two-step, liquid-solid extraction using 10 mL of 0.1 M EDTA-McIlviane buffer followed by 10 mL of methanol. Antibiotics recoveries from unprocessed manure, separated liquids, separated solids, and heat-treated solids using the two-step extraction method had relative standard deviations < 30% for all but ceftiofur. Total antibiotic recoveries were 67–131% for tetracyclines, 56% for sulfonamide, 49–53% for macrolides, and 1.3–66% for β-lactams. This is the first study to use one protocol to assess four classes of antibiotics in liquid and solid manure fractions. This study allowed for more precise risk assessment of antibiotic transport in manure waste stream applied to fields as a liquid or solid compost.

Western-style pork products have attracted many modern urban consumers, and these products have rapidly entered the Chinese market. The current hazard analysis of processed meat products mainly focuses on processing hazards (PAHs, microorganisms, and food additives), with less attention to veterinary drug residues. According to the survey results, the residues of antimicrobial drugs (sulfonamides and quinolones) in pork and its products in China are a severe problem, which may cause metabolic reactions, toxic effects, or enhance drug resistance. This study applied a modified QuEChERS method combined with ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MSMS) to develop a rapid and sensitive method for determining antimicrobial drugs in bacon and ham was successfully evaluated methodologically by EU 2002/657/EC. This study used a three-level, three-factor Box–Behnken design (BBD) to optimize the QuEChERS method by response surface methodology. The excellent linearity of the calibration curve was shown in the corresponding concentration range with a coefficient of determination greater than 0.99. The values of decision limit (CCα) and detection capability (CCβ) were in the range of 10.9–31.3 μg/kg and 11.8–52.5 μg/kg, respectively. The method successfully detected two trace levels of antimicrobial drugs in commercially available samples, including sulfadiazine and moxifloxacin.

Traditional Chinese medicine considers Lonicerae japonicae flos to have antibacterial detoxification, liver protection, and gallbladder protection. At present, studies have proven that Lonicerae japonicae flos has a good therapeutic effect on liver injury. Therefore, to confirm the clinical applicability of Lonicerae japonicae flos in the treatment of liver injury, we were the first to compare the pharmacokinetics of an oral ethanol extract of Lonicerae japonicae flos in normal rats and carbon tetrachloride-induced liver injury model rats. A method was developed for the simultaneous determination of 3-caffeoylquinic acid, 4-caffeoylquinic acid, 5-caffeoylquinic acid, 3,5-dicaffeoylquinic acid, 4,5-dicaffeoylquinic acid, protocatechuic acid, Sweroside, and Secoxyloganin in rat plasma by ultra-performance liquid chromatography tandem mass spectrometry. The results show that the method is reliable and reproducible and can be used for quantitative determination of biological samples. The pharmacokinetic parameters showed that the area under the concentration–time curve of eight compounds in the model group was significantly increased. The results showed that the total absorption of the active components of Lonicerae japonicae flos in the blood increased, the clearance rate slowed down, and the bioavailability of Lonicerae japonicae flos increased in liver injury diseases.

The global population is aging, and intervention strategies for anti-aging and the prevention of aging-related diseases have become a topic actively explored today. Nicotinamide adenine dinucleotide (NAD+) is an important molecule in the metabolic process, and its content in tissues and cells decreases with age. The supplementation of nicotinamide mononucleotide (NMN), an important intermediate and precursor of NAD+, has increased NAD+ levels, and its safety has been demonstrated in rodents and human studies. However, the high content of NMN in natural plants has not been fully explored as herbal medicines for drug development. Here, we identified that the leaf of Cinnamomum verum J. Presl (C. verum) was the highest NMN content among the Plant Extract Library (PEL) with food experience, using ultra-performance liquid chromatography–tandem mass spectrometry (UPLC-MS/MS). To validate this result, the extraction and quantitative analysis of bark, leaf, root, and stem of fresh C. verum was conducted. The results revealed that the bark had the highest NMN content in C. verum (0.471 mg/100 g). Our study shed light on the prospects of developing natural plants in the context of NMN as drugs for anti-aging and prevention of aging-related diseases. The future should focus on the development and application of C. verum pharmaceutical formulations.

With the abuse of antibiotics, bacterial antibiotic resistance is becoming a major public healthcare issue. Natural plants, especially traditional Chinese herbal medicines, which have antibacterial activity, are important sources for discovering potential bacteriostatic agents. This study aimed to develop a fast and reliable method for screening out antimicrobial compounds targeting the MRSA membrane from Psoralea corylifolia Linn. seed. A UPLC-MS/MS method was applied to identify the prenylated flavonoids in major fractions from the extracts of Psoralea corylifolia Linn. seed. The broth microdilution method was used to determine the minimum inhibitory concentrations (MICs) of different fractions and compounds. The morphological and ultrastructural changes of MRSA were determined by scanning electron microscopy (SEM). The membrane-targeting mechanism of the active ingredients was explored by membrane integrity assays, membrane fluidity assays, membrane potential assays, ATP, and ROS determination. We identified eight prenylated flavonoids in Psoralea corylifolia Linn. seed. The antibacterial activity and mechanism studies showed that this type of compound has a unique destructive effect on MRSA cell membranes and does not result in drug resistance. The results revealed that prenylated flavonoids in Psoralea corylifolia Linn. seeds are promising candidates for the development of novel antibiotic agents to combat MRSA-associated infections.

This study aims to simultaneously extract heterocyclic amines (HAs) and polycyclic aromatic hydrocarbons (PAHs) from ground pork for respective analysis by UPLC-MS/MS and GC-MS/MS, and study the effects of different flavorings and marinating time length on their formation and inhibition. Results showed that both HA and PAH contents followed a time-dependent increase during marinating, with HAs being more susceptible to formation than PAHs. The total HA contents in unmarinated pork and juice was, respectively, 61.58 and 139.26 ng/g, and rose to 2986.46 and 1792.07 ng/g after 24-h marinating, which can be attributed to the elevation of reducing sugar and creatinine contents. The total PAH contents in unmarinated pork and juice were, respectively, 34.56 and 26.84 ng/g, and increased to 55.93 and 44.16 ng/g after 24-h marinating, which can be due to the increment of PAH precursors such as benzaldehyde, 2-cyclohexene-1-one and trans,trans-2,4-decadienal. Incorporation of 0.5% (w/v) cinnamon powder or 0.5% (w/v) green tea powder was effective in inhibiting HA formation with the former showing a more pronounced effect for marinated pork, while the latter was for marinated juice. However, their addition was only effective in inhibiting PAH formation in marinated pork. Principle component analysis revealed the relationship between HA and PAH formation in ground pork and juice during marinating.

Background Accurately measuring plasma aldosterone concentration is difficult but meaningful for primary aldosteronism (PA) diagnosis. Methods In this study, we developed an ultra‐performance liquid chromatography‐tandem mass spectrometry (UPLC‐MS/MS) method for plasma aldosterone detection, evaluated its performance according to guidelines issued by CLSI, including detection limit, linearity, precision, and compared it with chemiluminescence immunoassay. Then, a reference range of plasma aldosterone in young people was established by using this method. Results The lower limit of quantitation (LOQ) was 10 pg/ml. The mean recovery rates of analyte added to serum were 100.07–102.05% in different concentrations. The linearity range was 20–2000 pg/ml. Inter‐assay CVs were 2.20–3.97% at aldosterone concentrations of 65.66–854.75 pg/ml. The regression equation of UPLC‐MS/MS (x) and chemiluminescence immunoassay (y) was y = 1.002x + 65.854 (r = 0.9456, n = 237). The reference range of plasma aldosterone detected by UPLC‐MS/MS was 11.30–363.82 pg/ml in young people in South China, and there was no statistically significant difference in plasma aldosterone concentration between two genders. Conclusion In conclusion, UPLC‐MS/MS can rapidly and accurately detect plasma aldosterone and is appropriate for clinical application., Left: performance characteristics of built UPLC‐MS/MS method for aldosterone detection; Right: Bland‐Altman plot for methodological comparison between UPLC‐MS/MS and CLIA.

Pueraria lobata and its variety P. lobata var. thomsonii are both traditional Chinese medicines that have high nutritional and medical value, whereas another variety, P. lobata var. montana has low nutritional and medicinal value and can cause ecological disasters. The material basis of different nutritional and medicinal values, which are caused by metabolite differences among these varieties, remains to be further clarified. Here, we performed ultra performance liquid chromatography-tandem mass spectrometry based widely targeted metabolome analysis on Pueraria lobata, P. lobata var. thomsonii, and P. lobata var. montana. Among them, a total of 614 metabolites were identified, and distinguished from each other using orthogonal partial least squares discriminant analysis. Our results suggest that the nutritional differences between P. lobata and its varieties can be explained by variations in the abundance of amino acids, nucleotides, saccharides, and lipids, differences in flavonoids, isoflavones, phenolic acids, organic acids, and coumarins contents caused the differences in the medicinal quality of P. lobata and its varieties. Additionally, the key metabolites responsible for the classification of the three Pueraria varieties were identified. This study provides new insights into the underlying metabolic causes of nutritional and medicinal variation in P. lobata and its varieties.

Propyl-propane-thiosulfonate (PTSO) and Propyl-propane-thiosulfinate (PTS) are organosulfur compounds used to supplement the diet of livestock because of their beneficial effects on feed palatability, their antibacterial, anti-inflammatory, and antimethanogenic activities. Besides, antibiotic residues in the environment can be reduced by using these natural bioactive compounds. The objective of this study was to optimize the extraction parameters for the analysis of PTSO and PTS in feed matrices by performing a solid-liquid extraction and quantification by Ultra performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS). Optimization was performed using the Response Surface Methodology on a Box–Behnken experimental design, optimizing the following parameters: solvent:sample ratios and evaporation temperature set for the rotary evaporator. The method was validated for 3 concentration levels for both PTSO (100, 500, 1000 ng g−1) and PTS (500, 1150, 2300 ng g−1). The highest recoveries of PTSO and PTS were obtained using 12.5 mL of 100% acetonitrile, stirring for 15 min, and an evaporation temperature of 20 °C. The validated method was further applied to detect and quantify these compounds in different feed matrices. In conclusion, this is the first study to simultaneously analyze PTSO and PTS at low concentrations, employing a sensitive technique such as UPLC-MS/MS. Ministerio de Ciencia, Innovación y Universidades (RTC-2017-6199-2) Junta de Andalucía (AT-17-5323)

Study of the occurrence and fate of thyroid hormones in aquatic environments requires a highly sensitive method that is capable of detecting such compounds at sub-ng/L concentrations. By using isotope-dilution UPLC-MS/MS, we developed a sensitive method for quantifying thyroxine (T4), 3,3ʹ,5-triiodothyronine (T3), 3,3ʹ,5ʹ-triiodothyronine (rT3), iodotyrosine (MIT), and 3,5-diiodotyrosine (DIT) in lake water and sediment samples. MIT and DIT were first reported in aquatic environments in this study and were detected in all sediment and water samples with concentrations of 0.01–1.1 ng/g dw and 1.9–9.9 ng/L, respectively. T4 was only observed in sediment, with the measured concentrations and frequency of

Background: Although newborn screening (NBS) for metabolic defects using the marker butyl carnitine (C4) combined with the C4-to-acetylcarnitine ratio is adequate, the incorporation of novel parameters may improve differential testing for these disorders without compromising sensitivity.Methods: Analytical and clinical performance was evaluated by MS/MS using 237 initially positive neonatal samples between March 2019 and March 2020 at the Newborn Screening Center of Xuzhou Maternity and Child Health Care Hospital. Additionally, second-tier testing by ultraperformance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) combined with the quantification of ethylmalonate (EMA) or isobutyryl-glycine (IBG) in dried blood spots (DBSs) was performed to reduce the false-positive rate.Results: We reviewed initial MS/MS data for DBSs from 469,730 neonates, and a second-tier test was performed using 237 samples that exceeded the C4 concentration cutoff value. Eleven variants of the ACADS gene were identified, with c.1031A>G (p.E344G) being the most common. Fifteen ACAD8 mutations were identified in seven patients, and Swiss modeling and amino acid conservation analyses were conducted for the novel variants. Based on a retrospective analysis of EMA and IBG, the application of second-tier tests before the release of neonatal screening results reduced referrals by over 91.89% and improved the positive predictive value (PPV) for short-chain acyl-CoA dehydrogenase deficiency/isobutyryl-CoA dehydrogenase deficiency (SCADD/IBDD) screening.Conclusion: A screening algorithm including EMA/IBG improves target differential testing for NBS and may eliminate unnecessary referrals while maintaining 100% sensitivity. Second-tier screening using UPLC-MS/MS as a rapid and convenient supplemental DNA sequencing method may be beneficial for differential detection.