Lignans from Opuntia ficus-indica seeds protect rat primary hepatocytes and HepG2 cells against ethanol-induced oxidative stress

Bioactivity-guided isolation of Opuntia ficus-indica (Cactaceae) seeds against ethanol-treated primary rat hepatocytes yielded six lignan compounds. Among the isolates, furofuran lignans 4?6, significantly protected rat hepatocytes against ethanol-induced oxidative stress by reducing intracellular reactive oxygen species levels, preserving antioxidative defense enzyme activities, and maintaining the glutathione content. Moreover, 4 dose-dependently induced the heme oxygenase-1 expression in HepG2 cells.     

Lignans in treatment of cancer and other diseases†

Lignans are widely distributed in nature and display an impressive range of biological activities. The extensive pharmaceutical use of lignans is linked to their antitumor, antiviral, hepatoprotective, and platelet activating factor (PAF) antagonistic activities, among many others. This review article highlights selected pharmacologically active lignans, outlines their therapeutic relevance to the treatment of cancer and other diseases, and accentuates research on plant-derived lignans, particularly preclinical lead identification and optimization studies performed in the authors' Natural Products Laboratory (NPL). Antitumor and anti-HIV lignans have been discovered and developed in the NPL by using bioactivity-directed fractionation and isolation as well as rational drug design-based structural modification and analog synthesis approaches. Notably, a significant and ongoing project on podophyllotoxin and its semisynthetic analog etoposide has led to the development of a potent derivative designated GL-331. GL-331 has received investigational new drug (IND) approval from the FDA and is currently in clinical trials against various cancers, especially drug-resistant cancers. Further design, synthesis, and evaluation of GL-331-related analogs are discussed.     

Hepatoprotective effect of total flavonoids from Laggera alata against carbon tetrachloride-induced injury in primary cultured neonatal rat hepatocytes and in rats with hepatic damage

Summary The hepatoprotective activities of total flavonoids of Laggera alata (TFLA) were evaluated by carbon tetrachloride (CCl₄)-induced injury in primary cultured neonatal rat hepatocytes and in rats with hepatic damage. In vitro, TFLA at a concentration range of 1–100 g/ml improved cell viability and inhibited cellular leakage of two enzymes, hepatocyte aspartate aminotransferase (AST) and alanine aminotransferase (ALT), caused by CCl₄. In vivo, oral treatment with TFLA at doses of 50, 100, and 200 mg/kg significantly reduced the levels of AST, ALT, total protein, and albumin in serum and the hydroxyproline and sialic acid levels in liver. Histopathological examinations revealed that liver damage were improved when treated with TFLA. Meanwhile, 1,1-diphenyl-2-picrylhydrazyl (DPPH) and superoxide radicals scavenging activities of TFLA were also determinated. To understand the exact components of TFLA responsible for the hepatoprotective effect, nine flavonoid compounds were isolated and identified from TFLA. In conclusion, the present investigation was the first to verify the hepatoprotective effect of L. alata in vitro and in vivo. The hepatoprotective action of TFLA is likely related to its potent antioxidative and anti-inflammatory activity. Neutralizing reactive oxygen species by nonenzymatic mechanisms and enhancing the activity of original natural hepatic-antioxidant enzymes may be the main mechanisms of TFLA against CCl₄-induced injury.     

Free radical scavenging and hepatoprotective actions of Quercus aliena acorn extract against CCl4-induced liver

In the present study, we investigated the protective effect of Quercus aliena acorn extracts against CCl₄-induced hepatotoxicity in rats, and the mechanism underlying the protective effects. Aqueous extracts of Quercus aliena acorn had higher superoxide radical scavenging activity than other types of extracts. The Quercus aliena acorn extracts displayed dose-dependent superoxide radical scavenging activity (IC₅₀ = 4.92 μg/ml), as assayed by the electron spin resonance (ESR) spin-trapping technique. Pretreatment with Quercus aliena acorn extracts reduced the increase in serum aspartate aminotransferase (AST) and serum alanine aminotransferase (ALT) levels. The hepatoprotective action was confirmed by histological observation. The aqueous extracts reversed CCl₄-induced liver injury and had an antioxidant action in assays of FeCl₂- ascorbic acid induced lipid peroxidation in rats. Expression of cytochrome P450 2E1 (CYP2E1) mRNA, as measured by RT-PCR, was significantly decreased in the livers of Quercus aliena acorn-pretreated rats compared with the livers of the control group. These results suggest that the hepatoprotective effects of Quercus aliena acorn extract are related to its antioxidative activity and effect on the expression of CYP2E1.     

Growth of microalgae Botryococcus sp. in domestic wastewater and application of statistical analysis for the optimization of flocculation using alum and chitosan

Microalga biomass has been recognized as a sustainable bio-product to replace terrestrial biomass in biofuel production. The microalga industry has high operating costs, specifically on harvesting and biomass recovery. Therefore, the development of an efficient harvesting method is crucial to the minimization of production cost. A statistical analysis through response surface methodology was used to investigate the optimization of harvesting efficiency using alum and chitosan as a coagulant. Growth rate and biomass productivity were also determined. This research revealed that the harvesting efficiency using alum was 99.3%, with optimum dosage and pH of 177.74 mg L^?1 and 8.24, respectively. Chitosan achieved 94.2% biomass recovery at an optimal dosage of 169.95 mg L^?1 at pH of 12. Moreover, Botryococcus sp. achieved the maximum growth of 0.7551 µ_max d^?1, with an average total biomass productivity of 9.81 mg L^?1?d^?1 in domestic wastewater. Overall, this study shows that both alum and chitosan coagulants have great potential for efficient microalgal biomass recovery. It suggests that domestic wastewater as a potential growth medium for the large-scale production of microalga biomass.     

Developing subunit immunogens using B and T cell epitopes and their constructs derived from the F1 antigen of Yersinia pestis using novel delivery vehicles

Yersinia pestis is the etiological agent of pneumonic and bubonic plague. As the currently licensed vaccines for plague have their own limitations, there is a need for a rational and more effective form of a subunit vaccine to combat both forms of the disease. Newer methods of antigen delivery coupled with adjuvant offer an alternative approach toward a plague vaccine. In order to develop a new generation vaccine against plague, we chose an immunodominant, outer membrane capsular protein, F1 of Y. pestis. The immunogenicity of the peptide sequences, predicted to possess B (three sequences, B1, B2 and B3) and T (two sequences, T1 and T2) cell determinants, was studied in a murine model with different genetic backgrounds, using alhydrogel and liposomes as delivery vehicles. All the peptide sequences are immunogenic in all mouse strains and showed primary and secondary immune response. B2 peptide was found to be most immunogenic, followed by B1 and B3 peptides. Chimeras made between B and T structures proved highly immunogenic and the antibody levels are comparable with native F1 antigen, thereby proving that T1 and T2 are helper sequences. Interestingly, the liposome mode of immunization was found to be more immunogenic and generated higher affinity antibodies than the alum-based preparation. Immunization using a mixture of all the peptides further proved B2 to be immunodominant. The IgG isotype profile showed predominance of IgG1, IgG2b followed by IgG2a for all the formulations irrespective of mode of antigen delivery. Lymphocyte proliferation of spleen cells primed in vivo with peptides, B-T conjugates and F1 antigen followed by in vitro stimulation with these antigens in soluble (medium) and particulate (liposome) form, showed dose-dependent stimulation of T cells, while B-T constructs showed a higher stimulation index, comparable to F1 antigen. The liposome mode of antigen presentation showed higher lymphoproliferation of spleen cells. Of all the peptides tested, T1 and T2 sequences showed the highest stimulation indices. The pattern of cytokine levels was in the following order: interferon-gamma>interleukin-2>interleukin-4. In vivo protective studies of the B-T conjugates revealed that B1T1 and a mixture of conjugates showed a survival rate of 10 days. Thus, the study highlights the importance of B and T cell epitopes as peptide-based immunogens, being a serious alternative for plague vaccine.     

Comparison of Chemical Constituents and Pharmacological Effects of Different Varieties of Chrysanthemum Flos in China

Chrysanthemum Flos is the prestigious traditional Chinese medicinal material and the popular health drink. This article comprehensively evaluated the chemical constituents, antioxidant activity, and hepatoprotective effects of 25 common chrysanthemum varieties in China. Firstly, we analyzed the chemical compositions of water extracts of chrysanthemum using UPLC/Q-TOF-MS, and identified 29 chemical components. The results displayed that chrysanthemum was rich in chemical constituents, but there were significant differences in the contents of four phenolic acids and five flavonoids among different varieties, and the coefficient of variation (CVs) ranged from 35.96 % to 114.62 %. Then, the antioxidant activities of different chrysanthemums were investigated, respectively via 1,1-diphenyl-2-picrylhydrazyl (DPPH), 2,2’-azinobis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS), and Ferric Reducing Antioxidant Power (FRAP) assays. The spectrum-effect relationships between nine main components and antioxidant activities were investigated to identify the antioxidant constitutes in chrysanthemums. Meanwhile, H₂O₂-induced hepatocyte injury testing showed wide variation in cultivar antioxidant capacity, with Tongchengju (TCJ) producing the best effect (90.32 %), followed by Chuju (CJ; 85.78 %). In addition, the hepatoprotective effects of 8 mainstream varieties were determined by the model of acute alcoholic liver injury. They protected liver from injury by affecting relevant liver function and antioxidant indexes. Huangshangongju (HSG) could decrease aspartate aminotransferase (AST) activity by 39.27 % in liver tissue; Hangju-Fubaiju (HJ-FBJ), Jinsihuangju (JSH), and Chuju (CJ) significantly decreased the malondialdehyde (MDA) content of liver tissue, which reduced by more than 40 %; Jinsihuangju (JSH) of used for tea could double the content of glutathione (GSH) and had the similar effect on superoxide dismutase (SOD) as the positive group, showing significant antioxidant capacity. Therefore, this study confirmed that chrysanthemums are potential resources as antioxidants, functional foods, and medicinal materials. Importantly, it may provide a scientific support for further development and utilization of chrysanthemum, and screen excellent varieties for different demands.     

Modification of API-2c and Its Reactive Site

The inhibitory activity of API-2c was reduced by carboxymethylation of methionine residues but not by chemical modifications of lysine and arginine residues. This indicates that API-2c requires methionine residue for inhibitory activity. Dissociation of API-2c-subtilisin BPN′ complex under mild conditions (pH-gradient dialysis) resulted in the conversion of API-2c to modified form (API-2C*).

The amino acid composition of API-2c did not change upon enzymatic modification. However, modified API-2c had two NH₂-terminal sequences (Asp-Ser-Pro- and Ile-Tyr-Asp-) and two COOH-termini (Phe and Met), and gave two bands on SDS-polyacrylamide gel disc electrophoresis after reduction of disulfide bonds by 2-mercaptoethanol. From these results, it is concluded that the Met-Ile bond located within a disulfide bridge in API-2c molecule is the reactive site for inhibition of subtilisin BPN′.