Artemisia argyi (A. argyi) is a Chinese herbal medicine in China. The main active components are volatile oils, flavonoids, and other compounds, which have various pharmacological activities. Methoxylated flavonoids are the main active ingredients in A. argyi. Flavonoid O-methyltransferase (FOMT) is a key enzyme in the O-methylation of flavonoids. In order to further understand the function and characteristics of FOMT proteins, this paper carried out the whole genome mining and identification of FOMT genes in A. argyi and performed phylogenetic, chromosomal localization, gene sequence characterization, subcellular localization prediction, protein structure, gene structure analysis, and expression pattern analysis. The results showed that a total of 83 FOMT genes were identified in the genome of A. argyi. The phylogenetic tree shows that FOMT genes are divided into two subgroups, CCoAOMT (caffeoyl CoA O-methyltransferase) subfamily (32 genes) and COMT (caffeic acid O-methyltransferase) subfamily (51 genes). Gene sequence analysis showed that the number of amino acids encoded by FOMT was 70-734 aa, the molecular weight was 25 296.55-34 241.3 Da, and the isoelectric point was 4.51-9.99. Compared with 32 members of the CCoAOMT subfamily, nearly 1/3 of the 51 members of the COMT subfamily were hydrophobic proteins and 2/3 were hydrophilic proteins. Subcellular localization prediction showed that more than 80% of CCoAOMT subfamily members were located in the cytoplasm, and 96% of COMT subfamily members were located in the chloroplast. COMT subfamily members have more motifs than CCoAOMT subfamily members. The N-terminal motifs of COMT subfamily proteins are relatively variable, while the C-terminal motifs are relatively conserved. Expression pattern analysis showed that CCoAOMT subfamily members were mainly expressed in roots, while COMT members were mainly expressed in leaves. Some FOMTs showed the tissue expression specificity by real-time quantitative PCR analysis, especially in leaves. In this study, we identified and analyzed the FOMT gene family in A. argyi, and provided a theoretical basis for further research on the function of FOMTs and the biosynthesis of methylated flavonoids in A. argyi.

The aim of this paper is to explore the key anti-fatigue active components in the saponin-like composition of American ginseng. The anti-fatigue activity of western ginseng samples was evaluated using a zebrafish model; metabolomics techniques were used to identify the main saponins in western ginseng from different origins; the active substances and relevant targets of the anti-fatigue effect of western ginseng were initially screened by constructing a PPI protein interaction network between western ginseng saponins and disease targets, and the key active ingredients were screened using a molecular docking method; finally, the anti-fatigue activity of the key active ingredients was evaluated using a zebrafish, animal experiment was approved by the Ethics Committee of Shandong Academy of Medical Sciences (SYXK20220005). The anti-fatigue activity of the key active ingredients was evaluated using a zebrafish model. The results of the zebrafish activity evaluation showed that there were significant differences in the activities of the western ginseng samples from the two origins, and a total of 10 different saponins were identified as possibly related to the anti-fatigue activity after further metabolomic testing and pattern discrimination. The core anti-fatigue targets were screened with the help of component-disease target PPI, combined with pharmacophore-like parameters and molecular docking techniques, and pseudoginsenoside F11 was found to have good binding activity to five of the targets. Finally, the zebrafish model revealed that pseudoginsenoside F11 exhibited significant anti-fatigue activity. This study used metabolomics and zebrafish model to screen the key active substances of pseudoginsenoside F11 for its anti-fatigue activity, which will provide a reference for further research on the anti-fatigue of pseudoginsenosides.

A new siderophore chelate (1) and 8 known compounds were identified from the liquid co-cultures of the marine-derived Streptomyces sp. IMB18-531 and Cladosporium sp. IMB19-099 by a combination of chromatography methods, including C18 reversed-phase medium pressure chromatography, gel column chromatography and HPLC. Their structures were determined by spectroscopic analysis and chemical methods as aluminioxamine E (1), desferrioxamine E (2), ferrioxamine E (3), terragine E (4), capsimicin (5), cyclo(L-prolinyl-L-tyrosine) (6), anthranilic acid (7), (Z)-14-methylpentadec-9-enoic acid (8), and (Z)-hexadec-8-enoic acid (9). Compound 2 showed inhibitory activities against the expression of liver fibrosis related genes COL1A1, MMP2, and TIMP2. Compounds 5, 8, and 9 displayed antibacterial activities against methicillin-resistant Staphylococcus aureus, S. epidermidis and Bacillus subtilis, with MICs of 16-64 μg·mL^-1. Compound 5 showed cytotoxicities against human pancreatic cancer MIA Paca-2 and human colon cancer HT-29 cell lines with IC₅₀ of 2.9 and 6.3 μmol·L^-1, respectively.

Ten dimeric phthalide racemates (1-10) were isolated from an aqueous extract of the Angelica sinensis root head (Guitou) by separation techniques of column chromatography over macroporous adsorbent resin, MCI resin, silica gel, and Sephadex LH-20, together with preparative thin-layer chromatography and reversed phase HPLC. The racemates were further separated into (+)-/(-)-1-(+)-/(-)-10 with chiral HPLC. Their structures including absolute configurations were elucidated by comprehensive analysis of spectroscopic data, combined with electronic circular dichroism (ECD) and NMR calculations as well as single crystal X-ray diffractions. Compounds (+)-/(-)-1-(+)-/(-)-10 are either new structure or new natural product, named (+)-/(-)-angelidipthalidic acids A-H [(+)-/(-)-1-(+)-/(-)-8] and (+)-/(-)-angelidipthalidols A and B [(+)-/(-)-9 and (+)-/(-)-10], respectively. Meanwhile, dimeric phthalide mono- and bis-lactone derivatives with 3.3′a, 8.6′- and 3.6′, 8.3′a-coupling patterns as well as determination of their relative configurations are discussed.

Mannitol-calcium chloride metal organic framework (MOF) cocrystal significantly improved the tabletability of β-mannitol and could be developed as a new tablet filler. However, mannitol monomer was found in the product during the scale-up production of the excipient, which significantly affected the functional properties of the excipient. In this study, we intend to quantify the multi-component eutectic system of mannitol-calcium chloride. In this experiment, the MOF cocrystal excipient mannitol-calcium chloride cocrystal was used as the model compound, and infrared spectrum was collected. Based on partial least squares regression (PLSR) method, the abnormal bands were removed and the spectrum was preprocessed by normalization. The quantitative correction model of mannitol-calcium chloride MOF cocrystal content in cocrystal excipients was established and compared by two different variable screening methods, genetic algorithm (GA) and competitive adaptive reweighting algorithm (CARS). Two different variable screening methods, GA method and CARS method, were used to screen out 160 and 14 variables, respectively. The mannitol-calcium chloride cocrystal model established by CARS-PLSR method had the best performance, and the average relative error (MRE) and corrected root mean square error (RMSEC) of the model were 0.008 8 and 0.892 5, respectively, the determination coefficient (R²) of the model was increased from 0.978 3 to 0.994 4. The quantitative method of eutectic system established in this study has high prediction accuracy, fast detection speed and good stability, which is of great significance for optimizing the preparation process conditions and quality control methods of such eutectic excipients.

With the development of small-molecule immunotherapy drugs, its combination with the programmed cell death ligand 1/programmed cell death protein 1 (PD-L1/PD-1) antibodies would provide a new opportunity for cancer treatment. Therefore, targeting PD-L1/PD-1 axis by small-molecule drug is an attractive approach to enhance antitumor immunity and considered as the next generation of tumor immunotherapy. In the present study, we investigated the anti-tumor role of salvianolic acid B (SAB) by regulating the PD-L1 level in tumors. Changes of total PD-L1 and membrane PD-L1 levels were determined by Western blot, flow cytometry and PD-1/PD-L1 interaction assays. The expression of mRNA level of PD-L1 was detected by real-time PCR. The cytotoxicity of activated peripheral blood mononuclear cell (PBMC) cells toward co-cultured tumor cells was measured by cell impedance assay and crystal violet experiment. Surface plasma resonance technique was used to analyze the direct interaction between SAB and ubiquitin carboxyl-terminal hydrolase 2 (USP2). The antitumor effect of SAB in vivo was examined by C57BL/6 mice bearing MC38 xenograft tumor (all animal experiments were conducted in accordance with the Animal Ethics Committee of the Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences). Western blot and flow cytometry assay showed that SAB can significantly downregulate the abundance of PD-L1 in RKO and PC3 cells in dose- and time-dependent manner. PD-1/PD-L1 binding assay revealed that SAB reduces the binding of tumor cells to recombinant PD-1 protein. Mechanism studies revealed that SAB can bind directly to USP2 protein and inhibit its activity, thus promote the ubiquitin-proteasome pathway degradation of PD-L1 proteins. In addition, Cell impedance and crystal violet staining indicated that SAB enhances the killing activity of co-cultured PBMC cells toward tumor cells. MC38 tumor transplanted mouse experiments revealed that SAB treatment displayed significant suppression in the growth of MC38 tumor xenografts in C57BL/6 mice with an inhibition rate of 63.2% at 20 mg·kg^-1. Our results demonstrate that SAB exerts its anti-tumor activity by direct binding and inhibiting the activity of USP2 and reducing the PD-L1 level. Our study provides an important material basis and scientific basis for the potential application of SAB in tumor immunotherapy drug targeting USP2-PD-L1 axis.

1-Deoxy-D-xylulose-5-phosphate synthase (DXS), the first key enzyme in 2-methyl-D-erythritol-4-phosphate (MEP) pathway, catalyzes the condensation of glyceraldehyde-3-phosphate with pyruvate to 1-deoxy-xylose-5-phosphate (DXP). In this study, PgDXS1, PgDXS2, and PgDXS3 genes were cloned from the root of Platycodon grandiflorum (P. grandiflorum). The open reading frame (ORF) of PgDXS1, PgDXS2, and PgDXS3 were 2 160, 2 208, and 2 151 bp in full length, encoding 719, 735, and 716 amino acids, respectively. Homologous alignment results showed a high identity of PgDXSs with DXS in Hevea brasiliensis, Datura stramonium and Stevia rebaudiana. The recombinant expression plasmids of pET-28a-PgDXSs were constructed and transformed into Escherichia coli (E. coli) BL21 (DE3) cells, and the induced proteins were successfully expressed. Subcellular localization results showed that PgDXS1 and PgDXS2 were mainly located in chloroplasts, and PgDXS3 was located in chloroplasts, nucleus and cytoplasm. The expression of three DXS genes in different tissues of two producing areas of P. grandiflorum were assayed via real-time fluorescence quantitative PCR, and the results showed that all of them were highly expressed in leaves of P. grandiflorum from Taihe. Under methyl jasmonate (MeJA) treatment, the expression levels of three PgDXS genes showed a trend of first decreasing and then increasing at different time points (3 - 48 h), and the activity of DXS showed a trend of first increasing and then decreasing in three tissues of P. grandiflorum. This study provides a reference for further elucidating the biological function of PgDXS in terpenoid synthesis pathway in P. grandiflorum.

Breast cancer has become the most prevalent malignant tumor among women, putting the health of women at serious risk. Screening for lead compounds in the active ingredients of plant that are effective and less toxic continues to be an important strategy for treating breast cancer. Gerbeloid J, a coumarin isolated from Gerbera piloselloides (L.) Cass., showed significant anti-cancer activity. But there is no report on the effect and mechanism of gerbeloid J on cycle and apoptosis of breast cancer. By using the CCK-8, clone formation, and PI staining assays, the effects of gerbeloid J on the proliferation of MCF-7 and MDA-MB-231 cells were assessed in this study. The effects of gerbeloid J on the apoptosis and mitochondrial function of MCF-7 and MDA-MB-231 cells were assessed using DAPI, Annexin V/TO-PRO-3, Rhod-2 AM, TMRM, DCFDA staining assays, and Western blot. The results demonstrated that gerbeloid J regulated the P21/CDC25C/CDK-1/cyclin B1 pathway and arrested the cell cycle at G2/M phase to suppressed the proliferation of MCF-7 and MDA-MB-231 cells. Additionally, gerbeloid J induced apoptosis through the stimulation of mitochondrial calcium excess, reduction of mitochondrial membrane potential, and promotion of ROS generation, and its mechanism was related to the activation of mitochondrial apoptotic pathway. In conclusion, by regulating the P21/CDC25C/CDK-1/cyclin B1 pathway and activating the mitochondrial apoptosis pathway, gerbeloid J could cause breast cancer cell cycle arrest and apoptosis, which might offer a promising candidate for the creation of new drugs against breast cancer.

Polymer nanomaterials have been attracted more and more attention because of their advantages such as long circulation, reduced immunogenicity and less side effects, and have become a hot research topic in nanomaterials. However, the number of polymer nanomedicines successfully applied in clinical application is very limited, and the unsatisfactory pharmacokinetic behavior is one of the main reasons for thisresult. After polymer nanoparticles enter the body, they will release free drugs and polymer excipients. Polymer nanoparticles are the loaded drugs and free drugs are the active chemicals for efficacy, while polymer excipients may cause excipient drug interactions. Therefore, the focus of the pharmacokinetics study of polymer nanoparticles should not be only limited to the free drugs themselves, but should also focus on the loaded drugs, free drugs and polymer excipients. The dynamic changes of polymer excipients and their metabolites pose new requirements and challenges for the bioanalysis of polymer nanomedicines. The characteristics and application scope of common analytical methods for detection polymer nanomedicines including chromatographic assay will be discussed in this paper. Moreover, this review will also summarize the absorption, distribution, metabolism and excretion of polymer nanomedicines. We hope this review will provide reference for the pharmacokinetics study, safety and effectiveness evaluation of polymer nanomedicines.

Liposome nanomedicine is a new drug preparation with nano scale, which is encapsulated by lipid bilayer vesicle structure. As a drug delivery carrier, liposome has many advantages such as good biocompatibility, biodegradation in vivo and strong targeting. The application of liposome nano drug delivery system can improve the pharmacokinetic behavior and efficacy of some drugs in vivo to a certain extent, and reduce toxic and side effects. After liposome nanomedicine enter into the body, free drugs will be released, so there will be loaded drugs and free drugs in the body. Loaded drugs are drug repositories, free drugs are related to their efficacy and adverse reactions. Therefore, the pharmacokinetics study of liposomes should focus on both loaded drugs and free drugs. Quantitative analysis of free drugs, liposome particles and their materials is a big challenge. The bioanalysis and pharmacokinetics of liposome nanomedicines will be introduced and discussed in this review. We hope this review will provide a reference for the development of liposome nanomedicine.