Unveiling the regulatory mechanisms of nodules development and quality formation in Panax notoginseng using multi-omics and MALDI-MSI.

INTRODUCTION: Renowned for its role in traditional Chinese medicine, Panax notoginseng exhibits healing properties including bidirectional regulatory effects on hematological system diseases. However, the presence of nodular structures near the top of the main root, known as nail heads, may impact the quality of the plant's valuable roots. OBJECTIVES: In this paper, we aim to systematically analyze nail heads to identify their potential correlation with P. notoginseng quality. Additionally, we will investigate the molecular mechanisms behind nail head development. METHODS: Morphological characteristics and anatomical features were analyzed to determine the biological properties of nail heads. Active component analysis and MALDI mass spectrometry imaging (MALDI-MSI) were performed to determine the correlation between nail heads and P. notoginseng quality. Phytohormone quantitation, MALDI-MSI, RNA-seq, and Arabidopsis transformation were conducted to elucidate the mechanisms of nail head formation. Finally, protein-nucleic acid and protein-protein interactions were investigated to construct a transcriptional regulatory network of nodule development and quality formation. RESULTS: Our analyses have revealed that nail heads originate from an undeveloped lateral root. The content of ginsenosides was found to be positively associated with the amount of nail heads. Ginsenoside Rb1 specifically accumulated in the cortex of nail heads, while IAA, tZR and JAs also showed highest accumulation in the nodule. RNA-seq analysis identified PnIAA14 and PnCYP735A1 as inhibitors of lateral root development. PnMYB31 and PnMYB78 were found to form binary complexes with PnbHLH31 to synergistically regulate the expression of PnIAA14, PnCYP735A1, PnSS, and PnFPS. CONCLUSION: Our study details the major biological properties of nodular structures in P. notoginseng and outlines their impact on the quality of the herb. It was also determined that PnMYB31- and PnMYB78-PnbHLH31 regulate phytohormones and ginsenosides accumulation, further affecting plant development and quality. This research provides insights for quality evaluation and clinical applications of P. notoginseng.

An efficient Pt@MXene platform for the analysis of small-molecule natural products.

Small-molecule (m/z<500) natural products have rich biological activity and significant application value thus need to be effectively detected. Surface-assisted laser desorption/ionization mass spectrometry (SALDI MS) has become a powerful detection tool for small-molecule analysis. However, more efficient substrates need to be developed to improve the efficiency of SALDI MS. Thus, platinum nanoparticle-decorated Ti3C2 MXene (Pt@MXene) was synthesized in this study as an ideal substrate for SALDI MS in positive ion mode and exhibited excellent performance for the high-throughput detection of small molecules. Compared with using MXene, GO, and CHCA matrix, a stronger signal peak intensity and wider molecular coverage was obtained using Pt@MXene in the detection of small-molecule natural products, with a lower background, excellent salt and protein tolerance, good repeatability, and high detection sensitivity. The Pt@MXene substrate was also successfully used to quantify target molecules in medicinal plants. The proposed method has potentially wide application.

Ti3C2(OH)x-assisted LDI-TOF-MS for the rapid analysis of natural small molecules.

The inhomogeneous distribution of co-crystallized analytes and the traditional organic matrices as well as the intensive background interference in the low molecular weight range hinder the application of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) in the analysis of small-molecular compounds. New two-dimensional material MXene (e.g., Ti3C2) exerts better hydrophilicity, homogeneity and repeatability, and higher laser desorption efficiency, as well as less background interference than traditional organic matrices and other nanomaterial matrices such as titanium oxide, graphene, and gold nanostructures. This study was aimed to design Ti3C2 matrix with abundant hydroxyls on its surface, enhance the stability of this hydroxyl-rich Ti3C2 (Ti3C2(OH)x), and evaluate the analytical performances of Ti3C2(OH)x-assisted laser desorption/ionization time-of-flight mass spectrometry (LDI-TOF-MS) for small-molecular natural compounds in complex samples. The developed Ti3C2(OH)x showed the distinct advantages such as minimum background interference, high peak intensity (~105), high salt (0.6 M) and protein (0.5 mg/mL) tolerance, good repeatability (relative standard deviation<20%), and good stability after eight months of storage. Ti3C2(OH)x-assisted LDI-TOF-MS analysis could be used to rapidly identify Artemisia annua (a world-famous traditional Chinese medicine) and quantify the contents of the main chemical ingredients (oxymatrine (OXY) and matrine) of Compound Kushen Injection (CKI). Interestingly, the content of OXY in CKI could be accurately quantified by Ti3C2(OH)x-assisted LDI-TOF-MS, and there was a good linear relationship (R2 -0.9929), a low limit of detection (400 pg), and a low limit of quantification (600 pg) of OXY. Taken together, the rapid and accurate analysis of small-molecular natural compounds in complicated samples could be achieved by the Ti3C2(OH)x-assisted LDI-TOF-MS analysis.

[Spatial distribution characteristics of metabolities in rhizome of Paris polyphylla var. yunnanensis: based on MALDI-MSI].

In this study, a method was established for in-situ visualization of metabolite distribution in the rhizome of Paris polyphylla var. yunnanensis. To be specific, through matrix-assisted laser desorption/ionization-mass spectrometry imaging(MALDI-MSI), the spatial locations of steroidal saponins, amino acids, organic acids, phytosterols, phytoecdysones, nucleosides, and esters in rhizome of the medicinal plant were directly analyzed, and six unknown compounds with differential distribution in rhizome tissues were identified. The specific procedure is as follows: preparation of rhizome tissue section, matrix screening and optimization, and MALDI-MSI analysis. The results showed that the steroidal saponins were mainly distributed in the central, amino acids in epidermis and cortex, low-molecular-weight organic acids in central epidermis, phytosterols in the epidermis and lateral cortex, the phytoecdysones in epidermis and cortex, nucleosides(uneven distribution) in epidermis and cortex, growth hormones around the epidermis and cortex, particularly outside the cortex, and esters in cortex with unobvious difference among different tissues. In this study, the spatial distribution of meta-bolites in the rhizome of P. polyphylla var. yunnanensis was characterized for the first time. The result can serve as a reference for identifying and extracting endogenous metabolites of P. polyphylla var. yunnanensis, exploring the synthesis and metabolism mechanisms of the metabolites, and evaluating the quality of medicinal materials.

High-throughput screening and spatial profiling of low-mass pesticides using a novel Ti3C2 MXene nanowire (TMN) as MALDI MS matrix.

Pesticides play critical roles in agricultural fields; however, pesticide residues can cause serious damage to human health and the ecological environment; therefore, developing a rapid and sensitive method for pesticide detection is urgently needed. Nanostructure-assisted matrix laser desorption/ionization (MALDI) mass spectrometry (MS) has great potential for the detection of low-mass pesticides. In this study, a novel Ti3C2 MXene nanowire (TMN) was prepared by a facile sol-gel method and served as a matrix to enhance MALDI MS performance in the analysis of pesticides in positive ion mode. The TMN showed superior performance in the high-throughput detection of six kinds of pesticides (organophosphorus, organochlorine, carbamate, neonicotinoids, triazole, and oxadiazines), with ultrahigh sensitivity (detection limits at sub-ppt levels), remarkable repeatability, excellent salt tolerance, and extremely low background compared to traditional organic matrices due to the specific polyaromatic structure and the doping of nitrogen. Furthermore, this matrix was successfully employed for the analysis of residual pesticides in traditional Chinese herbs, and the level of diniconazole was quantified with a linear range of 0-50 ng/mL (R2 > 0.99). More importantly, the spatial distribution of various endogenous compounds (e.g., amino acids and saccharides, fatty acids, alkaloids, and plant hormones) and xenobiotic pesticides from the intact root of the medicinal plant P. quinquefolium was clearly visualized using the TMN self-assembly film as a matrix for MALDI imaging mass spectrometry (IMS). With superior advantages such as sensitivity, simplicity, rapidness, and minimal sample requirement, TMN as a matrix-assisted MALDI MS shows great promise for various applications.

Development a multi-immunoaffinity column LC-MS-MS method for comprehensive investigation of mycotoxins contamination and co-occurrence in traditional Chinese medicinal materials.

Mycotoxins are poisonous secondary fungal toxic metabolites and harmful to human health. Traditional Chinese medicinal materials (TCMs), including more than two hundred functional foods, are vulnerably bred fungi, causing spoilage and multi-mycotoxins contamination. This study established a simultaneous analytical method by using multi-mycotoxins immunoaffinity column (multi-IAC) and HPLC-MS/MS to evaluate mycotoxins' contamination levels and natural incidence in TCMs. Aflatoxins (AFs, including AFB1, AFB2, AFG1 and AFG2), ochratoxin A (OTA), fumonisins (FB1 and FB2), zearalenone (ZEN), deoxynivalenol (DON) and T-2 toxins in three TCMs or functional foods of Polygalae Radix (PR), Coicis Semen (CS) and Eupolyphaga Steleophaga (ES) were detected. The systematically investigated results of 30 batch AFB1 positive samples revealed co-occurrence and correlation of multi-mycotoxins are significant differences in various matrices. All the samples in this study contain more than 5 mycotoxins. AFB1-AFs, AFB1-FBs, AFB1-DON, and AFB1-T-2 are the most observed co-occurrence, AFB1-OTA is also of concern due to its synergistic toxicity. This study's results can be used to establish guidelines for screening mycotoxin contaminants and limitations on acceptable levels in TCMs. Simultaneously, mycotoxin's correlation results in different matrices can also provide a reference for the standardization of TCM production and processing.

[Effect of 18-ß Glycyrrhetinic Acid on the Endoplasmic Reticulum of Nasal Epithelial Cells in Allergic Rhinitis Model Rats].

OBJECTIVE: To explore the effect of 18-ß glycyrrhetinic acid (GA) on the endoplasmic reticulum of nasal epithelial cells in allergic rhinitis (AR) model rats. METHODS: Totally 96 Wistar rats were randomly divided into the blank group, the AR model group, the loratadine group, the GA group, 24 in each group. AR models were established by peritoneally injecting ovalbumin (OVA). Morphological scoring was performed. GA at 21. 6 mg/kg was intragastrically administered to rats in the GA group. Nasal mucosal tissues were taken for electron microscopic examinations at the second, fourth, sixth, and tenth week after drug intervention. RESULTS: The overlapping score was 2.10 ± 0.45 in the blank group, 5.10 ± 0.56 in the loratadine group, 5.10 ± 0.56 in the AR model group, 5.20 ± 0.78 in the GA group, showing statistical difference when compared with the blank group (P < 0.01). Results under transmission electron microscope showed that the number of the endoplasmic reticulum increased in the AR model group, with obvious cystic dilatation, a lot of vacuole formation, and degranulation. A large number of free ribosomes could be seen in cytoplasm. With persistent allergen exposure, changes mentioned above was progressively aggravated in the endoplasmic reticulum of nasal mucosal epithelium in the AR model group. But the dilation of endoplasmic reticulum, vacuole formation, and degranulation were relieved in the GA group, and got close to those of the blank group. CONCLUSION: 18-ß GA could improve the expansion, vacuolization, and degranulation of the endoplasmic reticulum of nasal epithelial cells in AR model rats.