Citrus plants are economically and medicinally important crops worldwide. However, due to the highly complex genetic background of the genus, the accurate identification of medicinal species within this genus remains a major challenge. In this study, we employed the analysis of whole-GEnome (AGE), also known as the Shizhen Method, to conduct bioinformatics analysis on 49 genomes from 15 Citrus species, successfully constructing a candidate species-specific target sequence library that covers these species. The number of specific target sequences per species predominantly ranges from 10 000 to 100 000. From this candidate library, species-specific target sequences were selected for the corresponding species. Their high specificity was subsequently verified using Sanger sequencing and CRISPR-Cas12a technology, which confirmed their efficacy in identifying medicinal Citrus species. Based on this foundation, we established an AGE-based identification system for closely related Citrus species, which enabled the precise authentication of 30 commercial samples of Citri Reticulatae Semen and its decoction pieces. This study demonstrates that the AGE can be used for the accurate identification of medicinal Citrus species, which breaks through the bottlenecks of traditional methods and effectively resolves the identification challenges. It thereby provides technical support for quality control of Chinese herbal medicine and clinical medication safety.

The chemical constituents in the bark of Eucommia ulmoides were isolated and purified using chromatographic techniques, including silica gel, Sephadex LH-20, ODS, MCI, and semi-preparative HPLC. The structures of the compounds were elucidated by spectroscopic analyses (NMR, IR, UV, and MS). A total of six compounds were identified as duzhonglactone (1), (7S, 8R)-4-hydroxy-3, 3′, 5′-trimethoxy-8, 9-dinor-8, 4′-oxyneoligna-7, 9-diol-7′-aldehyde (2), armaosigenin (3), (-) evofolin B (4), N-trans-feruloyltyramine (5), and C-veratroylglycol (6). Among them, compound 1 was a new five-membered ring lactone, while compounds 2-5 were isolated for the first time from Eucommia ulmoides bark. Furthermore, compounds 2-4 demonstrated antioxidant activity, with IC₅₀ values for 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging of 46.34 ± 5.80, 74.17 ± 8.71, and 78.02 ± 8.94 μg·mL^-1, and for 2, 2′-azinobis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) scavenging of 50.56 ± 1.35, 54.37 ± 3.04, and 55.61 ± 3.41 μg·mL^-1, respectively.

This study aimed to investigate the interventional effect and potential mechanism of a lignan compound, (7S, 8R, 8'R)-eplignan C (EPC), from Ephedra intermedia, on ovalbumin (OVA)-sensitized asthmatic mice. Specific pathogen-free (SPF) male BALB/c mice were randomly assigned to normal control (CON), model (M), positive control (montelukast sodium, 1.3 mg·kg^-1), EPC low-dose (EPC-L, 10 mg·kg^-1), and EPC high-dose (EPC-H, 20 mg·kg^-1) groups. An allergic asthma model was established through intraperitoneal injection of OVA with aluminum hydroxide adjuvant followed by OVA aerosol challenge. After 7 consecutive days of administration, cough and asthma indicators, lung function, serum immunoglobulin E (IgE) levels, and lung tissue pathology were assessed 24 h post-final dose. Enzyme-linked immunosorbent assay (ELISA) was employed to measure serum levels of mast cell degranulation products and inflammatory factors; flow cytometry was utilized to detect immune cells (dendritic cells, mast cells, Tregs) and Ca²⁺ levels; immunofluorescence was applied to evaluate tryptase alpha/beta 1 (TPSAB1) and septin 11 (Septin11) protein expression in lung tissue; Western blot analysis was conducted to examine protein expression and activation levels of key molecules in the Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) signaling pathway in lung tissue samples. Furthermore, a co-culture system comprising mouse lung epithelial cells (MLE-12) and primary mouse splenocytes, combined with the JAK2/STAT3-specific inhibitor tyrphostin AG490, was employed to elucidate the mechanism underlying EPC's intervention in OVA-sensitized asthma. Results demonstrated that EPC alleviated asthma-related cough, inhibited airway hyperresponsiveness, improved lung pathological damage, modulated immune cell levels, suppressed inflammatory responses and mast cell degranulation, exhibiting significant anti-asthmatic effects, and reduced key protein expression in the JAK2/STAT3 signaling pathway. Further mechanistic investigation revealed that EPC regulated Th17/Treg immune imbalance and attenuated inflammatory responses in the co-culture system, effects that were blocked by AG490. Thus, it is speculated that EPC may exert its anti-asthmatic effects by regulating the JAK2/STAT3 signaling pathway to inhibit mast cell degranulation and modulate immune-inflammatory responses. This animal experiment gas been approved by the Animal Ethics Committe of Henan University of Chinese Medicine (approval number DWLL2018080003).

Gastrointestinal mucus constitutes a critical physiological barrier in the gastrointestinal tract. Its highly glycosylated mucin network effectively hinders the transport of particles and drug molecules toward the intestinal epithelium. In oral drug delivery, effectively overcoming this gastrointestinal mucus barrier is essential for achieving efficient drug absorption. On one hand, the continuously secreted mucus layer rapidly clears ineffectively penetrating particles due to its shear-thinning rheological property. On the other hand, macromolecular drugs face significant challenges in cross-mucus diffusion when their molecular size exceeds the mucus mesh pore size. Furthermore, certain small-molecule drugs can also be retained within the mucus layer through physicochemical interactions with its components, or rapidly eliminated by the dynamic turnover of the continuously secreted mucus, consequently leading to reduced drug delivery efficiency. Therefore, enhancing the mucus-penetrating capability of drug molecules is crucial for improving their oral bioavailability. This article systematically reviews the composition, structure, physicochemical properties, and barrier characteristics of gastrointestinal mucus, along with its barrier mechanisms affecting oral drug absorption. Additionally, it discusses nanotechnology-based drug delivery strategies designed to overcome the mucus barrier. The review aims to provide a theoretical basis and design principles for enhancing mucus permeability, improving oral bioavailability, and developing highly efficient mucus-penetrating nano-formulations.

The rhizosphere soil microenvironment critically influences secondary metabolite accumulation in medicinal rhizomatous plants. To compare the effects of the rhizosphere soil microenvironment in major production areas of Sichuan province on the accumulation of secondary metabolites in turmeric, we collected turmeric from Qianwei, Shuangliu, and Xingwen production areas in Sichuan province as research subjects. UPLC-Q-TOF-MS-based metabolomics was employed to analyze the composition of metabolites. And the high-throughput sequencing platform was utilized to analyze the rhizosphere soil microbial community structure. Then correlation analyses were performed between the metabolites, microbial community, and physicochemical factors. The results indicated that the metabolite profiles of turmeric from different production areas are relatively similar. The most abundant secondary metabolites in plants include ketones, aldehydes, esters, terpenoids, flavonoids, and polyphenols. Polyphenols, terpenoids, and some curcuminoids were significantly higher in turmeric from Shuangliu region than in samples from Qianwei and Xingwen. Significant differences were observed in rhizosphere soil physicochemical properties, including pH, organic matter, total nitrogen, available phosphorus, available potassium, manganese, magnesium, calcium, copper, and selenium among the different regions. At the phylum level, Proteobacteria and Acidobacteria were the dominant bacterial phyla across all three regions, while considerable variations existed at the genus level. In turmeric, correlation analysis revealed that bisdemethoxycurcumin correlated positively with pH, manganese, magnesium, and calcium. ar-Turmerone correlated positively with organic matter and available phosphorus. And tetrahydrocurcumin correlated positively with Rhodanobacter and Sphingomonas. In summary, secondary metabolites differed between turmeric from different regions. These metabolites exhibited correlations with both the physicochemical properties of the rhizosphere soil and the associated microbial communities. These findings provide a scientific basis for cultivation base selection and ecosystem-based cultivation practices.

In recent years, the synthetic cannabinoid N-(1-amino-3, 3-dimethyl-1-oxobutan-2-yl)-1-butyl-1H-indazole-3-carboxamide (ADB-BUTINACA) has attracted widespread attention due to its high affinity for the CB1 receptor and associated abuse risk; however, its toxicity mechanism remains poorly understood. This study aimed to systematically investigate the toxicological mechanisms of ADB-BUTINACA using an integrated approach. We established an acute rat exposure model (low and high doses: 0.1 and 1 mg·kg^-1) and employed untargeted metabolomics combined with molecular docking analysis to examine its effects on the urinary metabolome and toxicity pathways. Our analysis identified 52 significantly altered metabolites, including taurine and phenylacetaldehyde, and revealed six disturbed metabolic pathways, such as phenylalanine metabolism and one-carbon metabolism. Notably, choline and methionine were pinpointed as potential dose-dependent biomarkers. Molecular docking results demonstrated a strong binding affinity (-7.8 kcal·mol^-1) between ADB-BUTINACA and aldehyde dehydrogenase ALDH3, suggesting that the compound inhibits ALDH3 activity. This inhibition likely disrupts phenylalanine metabolism, leading to the accumulation of phenylacetaldehyde and the upregulation of phenylacetylglycine, thereby inducing oxidative stress and potential cardiovascular and neurotoxicity. Concurrently, the downregulation of methionine and choline indicated an impairment of one-carbon metabolism, which may further exacerbate oxidative stress. In conclusion, this study elucidates the mechanism by which ADB-BUTINACA exposure induces oxidative stress primarily through disrupting phenylalanine and one-carbon metabolism, utilizing non-targeted metabolomics and molecular docking. The identified biomarkers provide a crucial theoretical foundation for future research on the toxicity mechanisms, clinical detection, and risk assessment of synthetic cannabinoids. The experimental protocol complied with the welfare and ethical requirements for experimental animals and was approved by the Animal Ethics Committee of Zhejiang Chinese Medical University (Approval number: 20220718-11).

This study developed a sustained-release lipid liquid crystalline (LLC) system for leuprolide acetate (LA) and investigated the feasibility of using the lipase inhibitor orlistat (Orl) to modulate the matrix degradation and prolong the drug release period. The LA-LLC formulation was characterized by using polarized light microscopy (PLM), rheological analysis, and in vitro release studies to guide optimization. The impact of Orl on the sustained-release performance of LA-LLC was evaluated through in vitro degradation, in vitro release, and pharmacokinetic studies in rats. All animal protocols were approved by the Laboratory Animals Care and Use Committee of Yantai University (registration number is YTU20210319) (Yantai, China). Results showed that the LLC system utilizing glycerol dioleate (GDO) as the oil phase exhibited optimal sustained-release characteristics. The optimized LA-LLC formulation comprised soybean phosphatidylcholine, GDO, ethanol, and LA at a mass ratio of 44.5∶44.5∶10∶1. This gel exhibited a hexagonal phase structure by PLM. It demonstrated shear-thinning capacity characteristic of a pseudoplastic fluid. In vitro degradation and release results indicated that Orl significantly inhibited lipid degradation and prolonged drug release. Pharmacokinetic results in rats indicated that the addition of inhibitor Orl prolonged the release period from 42 days to 56 days compared to the LA-LLC group. This study demonstrates that the LA-LLC system, with its structural stability and controllable release, can effectively prolong drug release by employing a lipase inhibitor to regulate matrix degradation. The results of the study provide a novel strategy for the sustained delivery of peptide drugs.

Based on the ProTide technology, structural modification was performed on the 3′-hydroxyl group of 2′-deoxy-5-fluorouridine (FdUR) to design and synthesize a series of novel 5-FU phosphoramidate diester dual prodrugs (A1-A10). The structures of all compounds were confirmed by spectroscopic techniques, and their anti-hepatocellular carcinoma activity was evaluated using the MTT assay. The results indicated that several compounds exhibited significant anti-liver tumor activity. Among them, compound A6 showed the most potent activity with IC₅₀ values of 0.15 μmol·L^-1 (against HepG2 cells) and 0.34 μmol·L^-1 (against Huh7 cells), which were superior to the positive control 11b. It also demonstrated good selectivity towards normal cells. Mechanistic studies revealed that ³¹P NMR monitoring confirmed that A6 follows the classic ProTide activation pathway and can be hydrolyzed by carboxypeptidase Y to remove the carboxylate ester. In vitro metabolic stability studies indicated that A6 remained stable in rat plasma for up to 48 h, whereas it underwent rapid activation in liver homogenate. Further in vivo experiments confirmed that in a mouse orthotopic Huh7 xenograft model, A6 (45 mg·kg^-1, administered every 2 days) achieved a tumor inhibition rate of 55.84%, which was significantly higher than that of the sorafenib control group (38.98%), without causing significant changes in body weight. The animal protocols were approved by the Animal Ethics Committee of Capital Medical University (No. AEEI-2025-536). This study successfully developed a class of 5-FU dual prodrug candidates with significant anti-HCC activity. Among them, A6 exhibited excellent in vitro activity, metabolic stability, and in vivo antitumor efficacy. Its activation mechanism aligned with the designed pathway, providing a valuable lead compound and theoretical support for anti-HCC drug development.

Helicide and rosmarinic acid possess sleep-improving properties but suffer from poor aqueous solubility. This study aimed to construct a dual-drug co-loaded amorphous system. Compatibility and intermolecular interactions were predicted using Hansen solubility parameters and molecular docking. Co-evaporates were prepared via the solvent co-evaporation method. Polarized light microscopy, X-ray diffraction, and differential scanning calorimetry analyses confirmed the formation of a single-phase binary co-amorphous system. Fourier-transform infrared spectroscopy, ultraviolet spectroscopy, and nuclear magnetic resonance analyses revealed that hydrogen bonds formed between the aromatic ketone carbonyl group of helicide and the carboxylic acid group of rosmarinic acid, while π-π stacking interactions occurred between their aromatic rings, constituting a stable intermolecular network. Co-amorphization significantly enhanced the apparent solubility of both drugs in pure water by 4.7-fold and 5.1-fold, respectively. In vitro dissolution studies indicated an initial rapid release phase, followed by limitations due to gelation. All animal experiments were conducted in strict accordance with the regulations of the Animal Experiment Ethics Committee of Yunnan University of Chinese Medicine, with approval number: YNUTCM-XMSS-G-20250059. Pharmacodynamic evaluation in a zebrafish insomnia model demonstrated that the co-amorphous system at low and medium doses prolonged total sleep time, increased the number of sleep bouts and average bout length, and shortened sleep latency compared to individual drugs or physical mixture. A combination index (CI) < 0.7 confirmed a synergistic sleep-promoting effect. The solvent co-evaporation method proved efficient and straightforward for preparing the co-amorphous system. This co-amorphous strategy demonstrates significant advantages in improving aqueous solubility and achieving synergistic hypnotic efficacy.

This study aims to investigate the inhibitory effects of cryptotanshinone (CPT) on colorectal cancer (CRC) both in vitro and in vivo, and to explore its potential mechanism of action based on ferroptosis. CRC HCT116 and HCT8 cells were treated with different concentrations of CPT. Cell counting, colony formation assays, and Annexin V/PI staining revealed that CPT significantly inhibited CRC cell viability and proliferation and promoted apoptosis. A CRC patient-derived organoid (PDO) (in accordance with the regulations of the Nankai University Biomedical Ethics Committee approval number NKUIRB2025036) model was established, and cell counting, hematoxylin-eosin (HE) staining, and immunohistochemistry (IHC) detection of the tumor marker Ki67 confirmed that CPT suppressed PDO growth and malignant proliferation. A mouse xenograft tumor model was established (in compliance with the regulations of the Animal Ethics Committee of Tianjin University of Traditional Chinese Medicine, approval number: TCM-LAEC2025259K2011). Measurements of tumor volume and weight, HE staining, and Ki67 IHC demonstrated that CPT significantly inhibited tumor growth in vivo. Further studies found that a ferroptosis inhibitor could reverse the inhibitory effect of CPT on CRC cell proliferation, and CPT showed synergistic effects with ferroptosis inducers. Western blot and other detection results showed that CPT reduced the ratio of intracellular reduced glutathione (GSH) to oxidized glutathione (GSSG), and increased the levels of 4-hydroxynonenal (4-HNE) and the lipid peroxidation product malondialdehyde (MDA). Mechanistic studies indicated that CPT decreased glutathione peroxidase 4 (GPX4) mRNA levels and protein expression in a concentration-dependent manner. In conclusion, CPT can induce ferroptosis in CRC cells through GPX4, thereby inhibiting CRC cell proliferation and exerting anti-colorectal cancer effects.