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

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

Integrated Metabolome and Lipidome Strategy to Reveal the Action Pattern of Paclobutrazol, a Plant Growth Retardant, in Varying the Chemical Constituents of Platycodon Root.

Platycodon root, a medicinal food homology species which has been used in Asian countries for hundreds of years, is now widely cultivated in China. Treatment with paclobutrazol, a typical plant growth retardant, has raised uncertainties regarding the quality of Platycodon root, which have been rarely investigated. In the present study, metabolomic and lipidomic differences were revealed by ultra-high performance liquid chromatography coupled to ion mobility-quadrupole time of flight mass spectrometry (UPLC-IM-QTOF-MS). A significant decrease of platycodigenin-type saponins was observed in the paclobutrazol-treated sample. Carrying out a comprehensive quantitative analysis, the contents of total saponins and saccharides were determined to illustrate the mode of action of paclobutrazol on Platycodon root. This study demonstrated an exemplary research model in explaining how the exogenous matter influences the chemical properties of medicinal plants, and therefore might provide insights into the reasonable application of plant growth regulators.

Plant metabolomics for studying the effect of two insecticides on comprehensive constituents of Lonicerae Japonicae Flos.

Pesticides' overuse and misuse have been reported to induce ingredient variations in herbal medicine, which is now gaining attention in the medicinal field as a form of alternative medicine. To date, available studies on pesticide-induced ingredient variations of herbal medicine are limited only on a few compounds and remain most others unexamined. In this study, a plant metabolomics-based strategy was performed to systematically explore the effects of two frequently used insecticides on the comprehensive constituents of Lonicerae Japonicae Flos (LJF), the flower buds of Lonicera japonica Thunb. Field trials were designed on a cultivating plot of L. japonica with controls and treatments of imidacloprid (IMI) and compound flonicamid and acetamiprid (CFA). Unbiased metabolite profiling was conducted by ultra-high performance liquid chromatography/quadrupole-Orbitrap mass spectrometer. After data pretreatment by automatic extraction and screening, a data matrix of metabolite features was submitted for statistical analyses. Consequently, 29 metabolic markers, including chlorogenic acids, iridoids and organic acid-glucosides were obtained and characterized. The relative quantitative assay was subsequently performed to monitor their variations across flowering developments. This is the first study that systematically explored the insecticide-induced metabolite variations of LJF while taking into account the inherent variability of flowering development. The results were beneficial for holistic quality assessment of LJF and significant for guiding scientific use of pesticides in the large-scale cultivation.

High performance liquid chromatography-tandem mass spectrometry method for residue determination of 39 plant growth regulators in root and rhizome Chinese herbs.

A simple, rapid, and effective method based on salting-out extraction and LC-MS/MS techniques was developed for the determination of 39 plant growth regulator (PGR) residues in two of the most common root and rhizome Chinese herbs, Codonopsis Radix (Dangshen) and Notoginseng Radix et Rhizoma (Sanqi). The extraction process was performed with acetonitrile-water (5:1) and citrate buffer extraction salt. The performance of the method was validated in accordance with the analytical quality control criteria of SANTE/11813/2017 guidelines. Analyte recoveries of 79.49-109.41% (Dangshen) and 80.17-102.81% (Sanqi) were achieved. The limit of quantifications (LOQs) were determined with the consideration of accuracy and precision. LOQs were lower than the lowest residue limits in EU pesticide regulation (10 µg/kg) for most PGRs. Moreover, the method was successfully applied in the analysis of 35 batches of Dangshen, and 60 batches of Sanqi products. The concentration of eleven PGRs were determined in analyzed samples.

An integrated approach for global profiling of multi-type constituents: Comprehensive chemical characterization of Lonicerae Japonicae Flos as a case study.

To fully capture the chemo-diversity of medicinal plants is very essential for understanding of their pharmacological activities and guiding scientific quality control. Aiming to facilitate chemical characterization and novel natural products discovery, the present study proposed an integrated approach based on two-dimensional liquid chromatography coupled with quadrupole-Orbitrap mass spectrometer (2D LC/Q-Orbitrap MS). An offline comprehensive two-dimensional (2D) LC system was constructed to cover and separate multi-type constituents by combining hydrophilic interaction chromatography (HILIC) and conventional reversed phase C18. A two-step mass defect filtering-induced exclusion list-data dependent acquisition was developed to increase MS/MS coverage and selectivity. Additionally, an efficient interpretation strategy, combining an automatic matching algorithm and molecular networking (MN), was introduced for rapid recognition of known compounds and efficient elucidation of unreported ones. As a case study, the integrated approach was tentatively applied for comprehensive characterization of complex multi-type components in Lonicerae Japonicae Flos (LJF), a traditional Chinese medicine. Consequently, a total of 537 compounds were characterized from LJF, including a large number of potential novel structures. It was demonstrated that the integrated approach is powerful in deep investigation on chemical diversity of medicinal plants and discovery of novel structures. Its application could also be extended for global profiling of other complicated chemical systems, such as Chinese medicinal formulas.

Optimized QuEChERS Method Combined with UHPLC-MS/MS for the Simultaneous Determination of 15 Mycotoxins in Liquorice.

In our study, a reliable and rapid analytical method for the simultaneous determination of 15 mycotoxins (aflatoxin B1, aflatoxin B2, aflatoxin G1, aflatoxin G2, alternariol, agroclavine, citrinin, diacetoxyscirpenol, deoxynivalenol, fumonisin B1, fumonisin B2, ochratoxin A, sterigmatocystin, T-2 toxin, and zearalenone) in liquorice using ultra-HPLC coupled to tandem MS was developed and validated. Due to the complex ingredients in liquorice, we chose a QuEChERS-based extraction procedure as the sample pretreatment. Meanwhile, for the first time, acetate buffer was used to replaced water, which can greatly reduce the concentration of formic acid in acetonitrile, which further reduces the extraction efficiency of impurities. The optimal combination of adsorbents is 150 mg primary secondary amine, 150 mg silica gel, 600 mg octadecylsilane, and 900 mg anhydrous magnesium sulfate. Electrospray ionization in both positive- and negative-ionization modes was applied to detect all the mycotoxins in a single run time of 15 min, with LOQs in the range of 0.125-2.5 µg/kg. The recoveries of determination obtained were in the range of 81.0-104.7%, whereas the analytes could be accurately quantified in the 0.25-625 µg/kg concentration range, with all coefficients being >0.992. Intra- and interday reproducibility were lower than 5.5 and 8.9%, respectively, for all analytical mycotoxins. The validated method was finally applied to screen mycotoxins in 31 batches of real samples collected from drugstores and hospitals in Shanghai, China. Our survey findings show that six mycotoxins were detected, including alternariol, citrinin, deoxynivalenol, fumonisin B1, ochratoxin, and zearalenone, and that the positive rate of mycotoxins was 54.8% in real samples, ranging from 3.37 to 520.6 µg/kg.

Multi-mycotoxins analysis in Pheretima using ultra-high-performance liquid chromatography tandem mass spectrometry based on a modified QuEChERS method.

A sensitive and high-throughput method was established and validated for the simultaneous determination of 22 mycotoxins in Pheretima aspergillum (E.Perrier) and Pheretima guillelmi (Michaelsen). A modified Quick Easy Cheap Effective Rugged and Safe (QuEChERS) method was used for sample preparation with recoveries ranging from 73% to 105% with relative standard deviations (RSDs) <8.0% for all target analytes. Ultra-high-performance liquid chromatography coupled with triple quadrupole mass spectrometry (UHPLC-MS/MS) was applied for separation and detection in ESI (+) and ESI (-) modes with the limits of detection (LOD) in the range of 0.05-10µgkg-1. The 22 compounds could be accurately quantified in the 0.5-1000µgkg-1 concentration range with correlation coefficients >0.99. In all cases, the intra- and inter-day precisions were lower than 6% and 10%, respectively. Matrix-matched calibration was utilized for quantification purposes to compensate for the matrix effects. Furthermore, the established method was successfully applied in 17 batches of normal real samples collected from different areas of China and 2 batches of moldy samples due to improper storage, only mold-contaminated samples were confirmed to have fumonisin B1 (FB1) and fumonisin B2 (FB2) contamination at 2.54-3.78µgkg-1. The constructed method could serve as a practical application of the UHPLC-MS/MS method for the trace analysis of multiple mycotoxins in complex matrixes, especially for those with high lipid contents.

Multi-residue analysis of pesticides in traditional Chinese medicines using gas chromatography-negative chemical ionisation tandem mass spectrometry.

In this study, a residue analysis method for the simultaneous determination of 107 pesticides in traditional Chinese medicines (TCMs), Angelica sinensis, A. dahurica, Leonurus heterophyllus Sweet, Pogostemon cablin and Lonicera japonica Thunb., was developed using gas chromatography coupled with tandem mass spectrometry in negative chemical ionisation mode (GC-NCI-MS/MS). NCI has advantages of high sensitivity and selectivity to chemicals with electron-withdrawing groups, and yields low background interference. For sample preparation, QuEChERS (quick, easy, cheap, effective, rugged and safe) was applied. Due to the unique characteristics of TCMs, the clean-up step was optimised by adjusting amounts of primary secondary amine, C18, graphitised carbon black and silica sorbents. Validation was mainly performed by determining analyte recoveries at four different spiking concentrations of 10, 50, 100 and 200 ng g(-1), with seven replicates at each concentration. Method trueness, precision, linearity of calibration curves, lowest calibrated levels (LCLs) and matrix effects were determined to demonstrate method and instrument performance. Among the 107 pesticides tested, approximately 80% gave recoveries from 80% to 110% and < 10% relative standard deviation (RSD). The LCLs for nearly all pesticides were 5 ng g(-1), and as low as 0.1 ng g(-1) for dichlofenthion, endosulfan sulphate, flumetralin, isofenphos-methyl, methyl-pentachlorophenyl sulphide and trifluralin. The results indicate that GC-NCI-MS/MS is an excellent technique for quantitative and qualitative analysis of targeted GC-amenable pesticides at ultra-trace levels, especially in complex matrices such as TCMs.

[Determination of sulfur dioxide residues in sulfur fumigated Chinese herbs with headspace gas chromatography].

This paper aims to establish a method for the determination of sulfur dioxide in sulfur fumigation Chinese herbs. Sample powder and hydrochloric acid solution were isolated by paraffin layer in order to avoid early reactions, with the generation of sulfur dioxide, headspace with airtight needle was used to transfer sulfur dioxide into gas chromatograph, and detected with thermal conductivity detector. The analytical performance was demonstrated by the analysis of 12 herbs, spiked at four concentration levels. In general, the recoveries ranging from 70% to 110%, with relative standard deviations (RSDs) within 15%, were obtained. The limit of detection (LOD) was below 10 mg x kg(-1). Standard addition can be used for low recovery samples. The method is simple, less time-consuming, specific and sensitive. Methods comparison revealed that gas chromatography is better than traditional titration in terms of method operability, accuracy and specificity, showing good application value.

[Determination of seven toxaphene congeners in ginseng and milkvetch root by gas chromatography tandem mass spectrometry].

A novel method for the determination of representative toxaphene congeners in traditional Chinese herbal medicines was developed. Ginseng and Milkvetch Root were selected as the samples and seven toxaphene congeners were selected as the monitoring objects. The samples were extracted by accelerated solvent extraction with cyclohexane-acetone (9:1, v/v), then cleaned-up by Florisil solid phase extraction with hexane as the eluent and the residues were detected by gas chromatography-electron ionization tandem mass spectrometry (GC-EI-MS/MS) in multiple reaction monitoring (MRM) mode. The performance was demonstrated by the analysis of Ginseng and Milkvetch Root samples spiked with toxaphene congeners at three concentration levels of 0.005, 0.01 and 0.1 mg/kg. The recoveries ranged from 72.4% to 105% with the relative standard deviations (RSDs) of 0.96%-10.4%. The limits of detection (LODs) were 0.2-1.7 microg/kg. This method is sensitive and efficient in the aspect of extraction, and can be applied to monitor the residue of toxaphene congeners in Ginseng and Milkvetch Root.

[Simultaneous determination of 56 organochlorine and pyrethroid pesticides in traditional Chinese medicines by GC coupled with dual-tower and dual-column].

The paper is to report the establishment of a method for the determination of multi-residue organochlorine and pyrethroid pesticides in traditional Chinese medicines (TCMs). Fifty-six pesticides were extracted by high-speed homogenization, and then purified through gel permeation chromatography (GPC) and solid phase extraction (SPE) cartridges. The residues were simultaneously identified and quantified by GC-ECD equipped with dual tower, dual column and two micro-ECD detectors. The analytical performance was demonstrated by the analysis of 3 TCMs samples' extracts, spiked at three concentration levels for each pesticide. In general, the recoveries ranging from 70% to 110%, with relative standard deviations (RSDs) better than 15%, were obtained. The limit of detection (LOD) for most of the targeted pesticides tested was below 0.01 mg kg(-1). The method had good extraction efficiency, purification effect and good reproducibility, which could be applied to the determination of organochlorine and pyrethroid pesticide residues in the routine analysis of TCMs.