Inhibition of Liver Cancer HepG2 Cell Proliferation by Enzymatically Prepared Low-molecular Citrus Pectin.

BACKGROUND: Low-molecular citrus pectin (LCP) is a pectin polysaccharide with low molec-ular weight, low degree of crux, and no branching. It is obtained by degrading natural citrus pectin (CP) through physical, chemical and enzymatic methods. LCP has received considerable attention in recent years due to its potential applications in the medical and biological fields. METHODS: In our previous study, LCP was prepared from CP by using recombinant Bacillus subtilis pectate lyase B. Monosaccharide comparative analysis revealed that the galacturonic acid content of LCP was higher than that of CP. The cell viability effect of LCP was elucidated by using HepG2 cells and the Cell Counting Kit-8 (CCK-8) assay. Terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) staining, Annexin V-FITC/PI staining, and flow cytometer propidium iodide stain-ing were performed to detect the effects of LCP on apoptosis and cell cycle arrest in HepG2 cells. Mi-tochondrial membrane potential (MMP) was observed through 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethyl-imidacarbocyanine assay. RESULTS & DISCUSSION: The Mw of the prepared LCP was 7.6 kDa, which was significantly lower than that of CP (140 kDa). Cell viability decreased with the increase in the concentration of LCP. The half-inhibitory concentration of 1.46 ± 0.02 mg/mL was determined. Treatment with 1.6 mg/mL LCP in-duced the apoptosis of HepG2 cells with the inhibition rate of 83.10% ± 4.72%, and the cell cycle was arrested in the S phase. Furthermore, the MMP of HepG2 cells decreased with the increase in LCP concentration. CONCLUSION: The enzymatically prepared LCP could inhibit the proliferation of HepG2 cells. This study provided a partial experimental basis and reference for LCP to become a potential functional food for anti-liver cancer.

Combining simultaneous nitrification-endogenous denitrification and phosphorus removal with post-denitrification for low carbon/nitrogen wastewater treatment.

Due to the limited nutrient removal from low carbon/nitrogen (⩽4) wastewater, a process combined simultaneous nitrification-endogenous denitrification and phosphorus removal (SNDPR) with post-denitrification (PD) in a SBR was proposed for deep-level nutrient removal without external carbon addition. SNDPR driven by PAOs and GAOs reduced PO4(3-)-P (98.3%) and partial TN (59.0%) at low DO conditions (0.5±0.1mg/L), and post-dentrification achieved further NOX(-) (produced by SNDPR) removal (24.0%) anoxically by utilizing the residual intracellular polymers in GAOs. Combined control of anaerobic/aerobic/anoxic durations and low DO inhibition to aerobic GAOs and NOB conducted partial nitrification-endogenous denitrification (PNED) (66%), which saved 44.3% intracellular polymers to further reduce 64% TN in effluent. After 115-day operation, the average effluent PO4(3-)-P and TN concentrations were 0.4 and 3.9mg/L, respectively, with 92.1% of TN removal. Highly enriched PAOs (36%±2%), GAOs (22%±2%) and AOB (15%±3%) over NOB (3%±1%) facilitated P uptake, PNED and post-denitrification in the SNDPR-PD system.

A novel stoichiometries methodology to quantify functional microorganisms in simultaneous (partial) nitrification-endogenous denitrification and phosphorus removal (SNEDPR).

Although efficient removal of carbon (C), nitrogen (N) and phosphorous (P) from wastewater with low C/N ratio was achieved in anaerobic/aerobic simultaneous nitrification-endogenous denitrification and phosphorus removal (SNEDPR) systems, the removal pathways and metabolic transformations in this complex system are unclear. This work targeted at developing the stoichiometric models for denitrifying glycogen organisms (DGAOs) via nitrite and nitrate (DGAONi and DGAONa), and demonstrating a novel methodology to quantify diverse functional microorganisms (e.g. ammonia and nitrite oxidizing bacteria, aerobic phosphorus accumulating organisms (APAOs), denitrifying PAOs (DPAOs) and aerobic GAOs (AGAOs)) for the removal of C, N and P. The results showed that the anaerobic intracellular carbon storage (CODintra) was mainly accomplished by GAOs, and PAOs were only responsible for about 40% of CODintra through a stable P release. At the aerobic stage, 84.9% of P was removed by APAOs with 15.1% left by DPAOs, while 64.6% of N was removed by DGAOs (45.8% by DGAONi and 18.8% by DGAONa) with 18.1% by DPAOs and 17.3% by bacterial growth. High proportion of N removal via nitrite (partial nitrification-endogenous denitrification) (71%) saved 7.3% aeration and 38% intracellular carbon demand. However, AGAOs still activated well at the aerobic intercellular carbon consumption, which limited the further improvement of N removal efficiency. By elucidating the nutrient removal pathways among diverse functional microorganisms, the methodology developed in this study could accelerate the nutrient removal in the SNEDPR process.

[Experimental study on intervention effect of Grifola frondosa on nonalcoholic steatohepatitis].

To study the preventive effect of Grifola frondosa on nonalcoholic steatohepatitis (NASH). The rat model of NASH was established by feeding high-fat diets for 12 weeks and intervened with 0.5 g · kg(-1) · d(-1) and 1.0 g · kg(-1) · d(-1) of C. frondosa powder suspensions. The degrees of hepatocyte fatty degeneration and inflammation were observed under the optical microscope with routine HE staining. The NAFLD activity scores (NAS) were calculated. Serum ALT, AST and hepatic TG and CHOL were tested by the biochemical method. The hepatic MDA was examined by thiobarbituric acid method. The hepatic SOD was tested by the xanthine oxidase test. The hepatic GSH-PX activity was determined by the dithio-nitrobenzoic acid method. Hepatic TNF-α and IL-6 were detected by the enzyme-linked immunosorbent assay (ELISA). The NASH model group induced by high-fat diets showed higher hepatic NAS, ser- um ALT, AST, CHOL and hepatic TG, CHOL, MDA, TNF-α, IL-6 (P < 0.01 or P < 0.05) and lower serum TG and hepatic SOD, GSH-PX (P < 0.01, P < 0.05) than the normal control group. After being intervened with different doses of G. frondosa, the NASH group revealed significantly lower hepatic NAS, serum ALT and hepatic TG, CHOL, MDA, TNF-α and IL-6 (P < 0.05) and higher hepatic SOD, GSH-PX (P < 0.05) than the model group. G. frondosa may prevent the further development of NASH by improving the disorder of lipid metabolism in rats with NASH induced by high-fat diets, relieving the level of oxidative stress and reducing the generation of inflammatory cytokines.

Treating low carbon/nitrogen (C/N) wastewater in simultaneous nitrification-endogenous denitrification and phosphorous removal (SNDPR) systems by strengthening anaerobic intracellular carbon storage.

A novel simultaneous nitrification denitrification and phosphorous removal-sequencing batch reactor (SNDPR-SBR) enriched with PAOs (phosphorus accumulating organisms), DPAOs (denitrifying PAOs), and GAOs (glycogen accumulating organisms) at the ratio of 2:1:1 was developed to achieve the simultaneous nutrient and carbon removal treating domestic wastewater with low carbon/nitrogen ratio (≤3.5). The SNDPR system was operated for 120 days at extended anaerobic stage (3 h) and short aerobic stage at low oxygen concentration (2.5 h) with short sludge retention time (SRT) of 10.9 d and hydraulic retention time (HRT) of 14.6 h. The results showed that at the stable operating stage, the average effluent chemical oxygen demand (COD) and PO4(3-)-P concentrations were 47.2 and 0.2 mg L(-1), respectively, the total nitrogen (TN) removal efficiency was 77.7%, and the SND efficiency reached 49.3%. Extended anaerobic stage strengthened the intracellular carbon (mainly poly-ß-hydroxybutyrate, PHB) storage, efficiently utilized the organic substances in wastewater, and provided sufficient carbon sources for denitrification and phosphorus uptake without external carbon addition. Short aerobic stage at low oxygen concentration (dissolved oxygen (DO): 1 ± 0.3 mg L(-1)) achieved a concurrence of nitrification, endogenous denitrification, denitrifying and aerobic phosphorus uptake, and saved about 65% energy consumption for aeration. Microbial community analysis demonstrated that P removal was mainly performed by aerobic PAOs while N removal was mainly carried out by denitrifying GAOs (DGAOs), even though DPAOs were also participated in both N and P removal.