Isolating Antipathogenic Fungal Coumarins from Coriaria nepalensis and Determining Their Primary Mechanism In Vitro.

Through bioassay-guided isolation, eight undescribed coumarins (1-8), along with six reported coumarins (9-14), were obtained from Coriaria nepalensis. The new structures were determined by using IR, UV, NMR, HRESIMS, and ECD calculations. The results of the biological activity assays showed that compound 9 exhibited broad spectrum antifungal activities against all tested fungi in vitro and a significant inhibitory effect on Phytophthora nicotianae with an EC50 value of 3.00 µg/mL. Notably, compound 9 demonstrated greater curative and protective effects against tobacco balack shank than those of osthol in vivo. Thus, 9 was structurally modified to obtain new promising antifungal agents, and the novel derivatives (17b, 17j, and 17k) exhibited better effects on Sclerotinia sclerotiorum than did lead compound 9. Preliminary mechanistic exploration illustrated that 9 could enhance cell membrane permeability, destroy the morphology and ultrastructure of cells, and reduce the exopolysaccharide content of P. nicotianae mycelia. Furthermore, the cytotoxicity results revealed that compound 9 exhibited relatively low cytotoxicity against HEK293 cell lines with an inhibition rate of 33.54% at 30 µg/mL. This research is promising for the discovery of new fungicides from natural coumarins with satisfactory ecological compatibility.

Functional analyses of Toxoplasma gondii dihydroorotase reveal a promising anti-parasitic target.

Toxoplasma gondii relies heavily on the de novo pyrimidine biosynthesis pathway for fueling the high uridine-5'-monophosphate (UMP) demand during parasite growth. The third step of de novo pyrimidine biosynthesis is catalyzed by dihydroorotase (DHO), a metalloenzyme that catalyzes the reversible condensation of carbamoyl aspartate to dihydroorotate. Here, functional analyses of TgDHO reveal that tachyzoites lacking DHO are impaired in overall growth due to decreased levels of UMP, and the noticeably growth restriction could be partially rescued after supplementation with uracil or high concentrations of L-dihydroorotate in vitro. When pyrimidine salvage pathway is disrupted, both DHOH35A and DHOD284E mutant strains proliferated much slower than DHO-expressing parasites, suggesting an essential role of both TgDHO His35 and Asp284 residues in parasite growth. Additionally, DHO deletion causes the limitation of bradyzoite growth under the condition of uracil supplementation or uracil deprivation. During the infection in mice, the DHO-deficient parasites are avirulent, despite the generation of smaller tissue cysts. The results reveal that TgDHO contributes to parasite growth both in vitro and in vivo. The significantly differences between TgDHO and mammalian DHO reflect that DHO can be exploited to produce specific inhibitors targeting apicomplexan parasites. Moreover, potential DHO inhibitors exert beneficial effects on enzymatic activity of TgDHO and T. gondii growth in vitro. In conclusion, these data highlight the important role of TgDHO in parasite growth and reveal that it is a promising anti-parasitic target for future control of toxoplasmosis.

Four new pregnane C21 steroids isolated from the roots of Cynanchum auriculatum.

Four new steroids cynansteroid G-I (1-3) and cynansteroid K (4), a new natural product 5,6-deacidizingcaudatin (5), and a known compound glycocaudatin (6), were isolated from the roots of Cynanchum auriculatum. The structures of new compounds were identified by comprehensive spectroscopic analyses, including NMR, HRESI-MS, ECD, UV, and IR spectral data. The cytotoxic activities of all the isolates against two human tumour cell lines (COLO-205 and BGC-823) were screened, unfortunately, which were weaker than positive control.

The determinants regulating Toxoplasma gondii bradyzoite development.

Toxoplasma gondii is an obligate intracellular zoonotic pathogen capable of infecting almost all cells of warm-blooded vertebrates. In intermediate hosts, this parasite reproduces asexually in two forms, the tachyzoite form during acute infection that proliferates rapidly and the bradyzoite form during chronic infection that grows slowly. Depending on the growth condition, the two forms can interconvert. The conversion of tachyzoites to bradyzoites is critical for T. gondii transmission, and the reactivation of persistent bradyzoites in intermediate hosts may lead to symptomatic toxoplasmosis. However, the mechanisms that control bradyzoite differentiation have not been well studied. Here, we review recent advances in the study of bradyzoite biology and stage conversion, aiming to highlight the determinants associated with bradyzoite development and provide insights to design better strategies for controlling toxoplasmosis.

In vitro Anti-parasitic Activity of Pelargonium X. asperum Essential Oil Against Toxoplasma gondii.

Toxoplasmosis is a global zoonotic disease, and one-third of the human population is chronically infected by Toxoplasma gondii. Due to the limited effectiveness and prominent side effects of the existing drugs, there is a dire need for the discovery of new therapeutic options in the treatment of toxoplasmosis. In this study, five essential oils (EO) were screened for their anti-parasitic activity against T. gondii. The cytotoxicity of essential oils was evaluated using the MTT assay on human foreskin fibroblast cells. The CC50 values of Eucalyptus globulus EO, Cupressus sempervirens EO, Citrus aurantifolia EO, Melaleuca alternifolia EO, and Pelargonium X. asperum (Pa) EO were found to be 22.74, 7.25, 15.01, 6.26, and 4.77 mg/mL, respectively. Only PaEO exhibited anti-parasitic activity, and inhibited the growth of T. gondii in a dose-dependent manner. In addition, treatment with PaEO, was found to reduce the volume of T. gondii tachyzoites and make their membrane surfaces rough. These results showed that PaEO was able to inhibit the growth of T. gondii by reducing invasion, which may be due to its detrimental effect on the ability of tachyzoites to move. These findings suggest that PaEO could be a potential anti-T. gondii drug, which may facilitate the development of new and effective treatments against toxoplasmosis.

Garmultins A-G, Biogenetically Related Polycyclic Acylphloroglucinols from Garcinia multiflora.

Garmultins A and B (1 and 2), two polycyclic polyprenylated acylphloroglucinols characterized by the coupling of two novel cages, 2,11-dioxatricyclo[4.4.1.03,9]undecane and tricyclo[4.3.1.03,7]decane, along with five biogenetically related analogues (3-7), were isolated from Garcinia multiflora. Their structures and absolute configurations were determined by extensive NMR analysis, X-ray crystallography, and electronic circular dichroism calculations. Three compounds were capable of inhibiting oncogene expression and inducing apoptosis in human erythroleukemia cells.

Effect of artesunate supplementation on bacterial translocation and dysbiosis of gut microbiota in rats with liver cirrhosis.

AIM: To evaluate the effect of artesunate (AS) supplementation on bacterial translocation (BT) and gut microbiota in a rat model of liver cirrhosis. METHODS: Fifty-four male Sprague-Dawley rats were randomly divided into a normal control group (N), a liver cirrhosis group (M) and a liver cirrhosis group intervened with AS (MA). Each group was sampled at 4, 6 and 8 wk. Liver cirrhosis was induced by injection of carbon tetrachloride (CCl4), intragastric administration of 10% ethanol, and feeding a high fat diet. Rats in the MA group were intragastrically administered with AS (25 mg/kg body weight, once daily). Injuries of the liver and intestinal mucosa were assessed by hematoxylin-eosin or Masson's trichrome staining. Liver index was calculated as a ratio of the organ weight (g) to body weight (g). The gut microbiota was examined by automated ribosomal intergenic-spacer analysis of fecal DNA. BT was assessed by standard microbiological techniques in the blood, mesenteric lymph nodes (MLNs), liver, spleen, and kidney. RESULTS: Compared to group N, the body weight was reduced significantly in groups M and MA due to the development of liver cirrhosis over the period of 8 wk. The body weight was higher in group MA than in group M. The liver indices were significantly elevated at 4, 6 and 8 wk in groups M and MA compared to group N. AS supplementation partially decreased the liver indices in group MA. Marked histopathologic changes in the liver and small intestinal mucosa in group M were observed, which were alleviated in group MA. Levels of pro-inflammatory interleukin-6 and tumor necrosis factor-α were significantly elevated at 8 wk in ileal homogenates in group M compared to group N, which were decreased after AS supplementation in group MA. The dysbiosis of gut microbiota indicated by the mean diversity (Shannon index) and mean similarity (Sorenson index) was severe as the liver cirrhosis developed, and AS supplementation had an apparent intervention effect on the dysbiosis of gut microbiota at 4 wk. The occurrence of BT was increased in the liver of group M compared to that of group N. AS supplementation reduced BT in group MA at 8 wk. BT also occurred in the MLNs, spleen, and kidney, which was reduced by AS supplementation. BT was not detected in the blood in any group. CONCLUSION: Dysbiosis of gut microbiota, injury of intestinal mucosal barrier and BT occurred as liver cirrhosis progressed, which might enhance inflammation and aggravate liver injury. AS may have other non-antimalarial effects that modulate gut microbiota, inhibit BT and alleviate inflammation, resulting in a reduction in CCl4, alcohol and high fat-caused damages to the liver and intestine.

Diterpene alkaloids and diterpenes from Spiraea japonica and their anti-tobacco mosaic virus activity.

Five new naturally occurring natural products, including two atisine-type diterpene alkaloids (1 and 2), two atisane-type diterpenes (3 and 4), and a new natural product spiramine C2 (5), along with nine known ones (6-14), were isolated from the ethanolic extracts of the whole plant of Spiraea japonica var. acuminata Franch. Their structures were elucidated by extensive spectroscopic analysis. The anti-tobacco mosaic virus (TMV) activities of all the compounds were evaluated by the conventional half-leaf method. Six compounds (2, 3, 6, 7, 11, and 12) exhibited moderate activities at 100 µg/mL with inhibition rates in the range of 69.4-92.9%, which were higher than that of the positive control, ningnanmycin. Their preliminary structure-activity relationships were also discussed.

Limonoids from Munronia henryi and their anti-tobacco mosaic virus activity.

Five new compounds (1-5), including three limonoids, one diterpenoid, and one phytosterol, along with four known limonoids (6-9), were isolated from the ethanolic extracts of whole plants of Munronia henryi. Their structures were elucidated by extensive spectroscopic analysis. In addition, the anti-tobacco mosaic virus (TMV) activities of all the isolated compounds were evaluated by the conventional half-leaf and leaf-disk methods along with Western blot analysis. Most of the tested compounds showed strong antiviral activities, with IC50 values in the range of 14.8-34.0µg/mL, compared with ningnanmycin as a positive control (IC50=44.6µg/mL), and their preliminary structure-activity relationships were also discussed.

Two Unusual Polycyclic Polyprenylated Acylphloroglucinols, Including a Pair of Enantiomers from Garcinia multiflora.

Two polycyclic polyprenylated acylphloroglucinols, garcimulins A and B ((±)-1 and 2), including a pair of enantiomers with the unique caged tetracyclo[5.4.1.1(1,5).0(9,13)]tridecane skeleton were isolated from Garcinia multiflora. Their structures and absolute configurations were determined by extensive analysis of spectroscopic data and electronic circular dichroism (ECD) calculations. Compounds 1 and 2 exhibited cytotoxic activities against five human cancer cell lines in vitro (IC50 3.42-13.23 µM). The acidification of lysosomes in HeLa cell was obviously affected by compound 2.