Effect of particulate organic matter fractions on the distribution of heavy metals with aided phytostabilization at a zinc smelting waste slag site.

Particulate organic matter (POM) significantly affects the distribution of heavy metals in contaminated soil. However, the effect of POM on the fate of heavy metals during in situ-aided phytostabilization of waste slag is unclear. The objective of this study was to investigate the distributions of heavy metals such as Cu, Pb, Zn, and Cd in the POM fractions at a zinc smelting waste slag site under in situ-aided phytostabilization after five years. The results showed that the litters and residues of four plants-Arundo donax, Broussonetia papyrifera, Cryptomeria fortunei, and Robinia pseudoacacia-decomposed to form different POM size fractions. The percentage of the 0.05-0.25 mm POM size fraction was the highest, followed by the >1 mm and 0.5-1 mm POM size fractions, and that of the 0.25-0.5 mm POM size fraction was the lowest. The masses of POM derived from the four plants were in the following order: C. fortunei > B. papyrifera > A. donax > R. pseudoacacia. The contents, enrichment coefficients, and mass loads of heavy metals such as Cu, Pb, Zn, and Cd in the POM increased with decreasing POM size, and those in the 0.05-0.25 mm POM size fraction were the highest. The mass load of heavy metals in the POM occurred in the following order: Cu > Cd > Zn > Pb. The surfaces of the POM with coarser and smaller size fractions were smoother and rougher, respectively, and the smaller POM size fractions had larger specific surface areas. The main functional groups in the different POM size fractions were -COOH, -OH, CO, CC, C-H, Si-O, and -CH3. The POM fractions played a significant role in determining the distribution of heavy metals in the revegetated waste slag. These findings have important implications for aided phytostabilization, which significantly influences the fate and speciation of heavy metals at the phytoremediation site.

Molecular Docking and Molecular Dynamics Studies on Selective Synthesis of α-Amyrin and ß-Amyrin by Oxidosqualene Cyclases from Ilex Asprella.

Amyrins are the immediate precursors of many pharmaceutically important pentacyclic triterpenoids. Although various amyrin synthases have been identified, little is known about the relationship between protein structures and the constituent and content of the products. IaAS1 and IaAS2 identified from Ilex asprella in our previous work belong to multifunctional oxidosqualene cyclases and can produce α-amyrin and ß-amyrin at different ratios. More than 80% of total production of IaAS1 is α-amyrin; while IaAS2 mainly produces ß-amyrin with a yield of 95%. Here, we present a molecular modeling approach to explore the underlying mechanism for selective synthesis. The structures of IaAS1 and IaAS2 were constructed by homology modeling, and were evaluated by Ramachandran Plot and Verify 3D program. The enzyme-product conformations generated by molecular docking indicated that ASP484 residue plays an important role in the catalytic process; and TRP611 residue of IaAS2 had interaction with ß-amyrin through π-σ interaction. MM/GBSA binding free energy calculations and free energy decomposition after 50 ns molecular dynamics simulations were performed. The binding affinity between the main product and corresponding enzyme was higher than that of the by-product. Conserved amino acid residues such as TRP257; TYR259; PHE47; TRP534; TRP612; and TYR728 for IaAS1 (TRP257; TYR259; PHE473; TRP533; TRP611; and TYR727 for IaAS2) had strong interactions with both products. GLN450 and LYS372 had negative contribution to binding affinity between α-amyrin or ß-amyrin and IaAS1. LYS372 and ARG261 had strong repulsive effects for the binding of α-amyrin with IaAS2. The importance of Lys372 and TRP612 of IaAS1, and Lys372 and TRP611 of IaAS2, for synthesizing amyrins were confirmed by site-directed mutagenesis. The different patterns of residue-product interactions is the cause for the difference in the yields of two products.

Dioscin suppresses hepatocellular carcinoma tumor growth by inducing apoptosis and regulation of TP53, BAX, BCL2 and cleaved CASP3.

BACKGROUND: Hepatocellular carcinoma (HCC) is the most commonly diagnosed malignancy of the liver, occurs frequently in the setting of chronic liver injury. Although multiple therapeutic approaches are available, the prognosis of patients with HCC remains poor. Dioscin is a natural steroid saponin that presents in various plants. The anti-cancer and anti-fibrotic effects have been extensively reported. However, the effect of dioscin on HCC remains unclear. We aimed to investigate the anti-HCC properties of dioscin in vitro and in vivo. METHODS: MTT (3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyl-tetrazolium bromide) assay was used to analyze the growth inhibition activity of Dioscin in human cell lines, Bel-7402, HepG2, Lovo, and EAhy926. Antitumor activity through induction of apoptosis was evaluated by flow cytometry using Annexin-V and propidium iodide (PI) staining, laser scanning confocal microscopy (LSCM) analysis with Hochest33342 and PI labeling, and DNA fragmentation analysis. The expression of apoptosis-related proteins tumor protein p53 (TP53), BCL2-associated X protein (BAX), B-Cell CLL/Lymphoma 2 (BCL2) and Caspase 3 (CASP3) was measured by Western blot. Nude mice bearing Bel-7402 were administered intraperitoneally at different doses of dioscin and 5-FU (5-Fluorouracil) treatment was used as a control. Tumor volume and tumor weight of each mouse were then measured. RESULTS: We demonstrated that Dioscin inhibited proliferation of HCC cell lines in a dose-dependent manner. Dioscin also significantly induced morphological changes during death by apoptosis and increased DNA damage of Bel-7402 cells. Moreover, we demonstrated that Dioscin displayed anticancer activity via up-regulating expression of TP53, BAX and CASP3 protein, as well as down-regulating BCL2 in Bel-7402 cells. Notably, the in vivo anticancer activity of Dioscin was further assessed and achieved greater inhibition efficiency at the concentration increased to 24mg/kg/day than 5-FU at dose of 10mg/kg/day in nude mice bearing Bel-7402 cells. CONCLUSIONS: Dioscin inhibited tumor growth via inducing apoptosis, which was accompanied by altered expression of apoptotic pathway proteins, such as TP53, BAX, BCL2 and CASP3. Our findings indicate that further evaluation of dioscin as a novel therapeutic approach for HCC is warranted.