Hydroethanolic Extracts of Haplopappus baylahuen Remy and Aloysia citriodora Palau Have Bactericide Activity and Inhibit the Ability of Salmonella Enteritidis to Form Biofilm and Adhere to Human Intestinal Cells.

We analysed whether the hydroethanolic extracts from leaves of Haplopappus baylahuen Remy (bailahuen) and Aloysia citriodora Palau (cedron) inhibit the growth and ability of Salmonella Enteritidis to form biofilms and to adhere to human intestinal epithelial cells. Herein, we first determined the total phenolic content and antioxidant and antibacterial activities of the extracts. Then, Salmonella Enteritidis was treated with the extracts to analyse biofilm formation by scanning electronic microscopy and the violet crystal test. We also measured the efflux pump activity of Salmonella Enteritidis since biofilm formation is associated with this phenomenon. Furthermore, the human intestinal cell line Caco-2 was infected with Salmonella Enteritidis pretreated with the extracts, and 30 min later, the number of bacteria that adhered to the cell surface was quantified. Finally, we determined by qPCR the expression of genes associated with biofilm formation, namely, the diguanilate cyclase AdrA protein gene (adrA) and the BapA protein gene (bapA), and genes associated with adhesion, namely, the transcriptional regulator HilA (hilA). The phenolic content and antioxidant and bactericide activities were higher in bailahuen than in the cedron extract. Biofilm formation was inhibited by the extracts in a dose-dependent manner, while the activity of efflux pumps was decreased only with the cedron extract. Adhesion to Caco-2 cells was also inhibited without differences between doses and extracts. The extracts decreased the expression of adrA; with the cedron extract being the most efficient. The expression of hilA is affected only with the cedron extract. We concluded that hydroethanolic extracts of bailahuen and cedron differentially inhibit the growth of Salmonella Enteritidis and affect its the ability to form biofilms and to adhere to human intestinal epithelial cells. These results highlight the presence of molecules in bailahuen and cedron with a high potential for the control of the Salmonella Enteritidis pathogenesis.

Ethanol extract of Liriodendron chinense (Hemsl.) Sarg barks attenuates hyperuricemic nephropathy by inhibiting renal fibrosis and inflammation in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Liriodendron chinense (Hemsl.) Sarg, known as the Chinese tulip tree, has a long history of cultivation and utilization in many Asia countries, especially in China to use in traditional Chinese medicine for expelling "wind and dampness", a term corresponding to rheumatic fever and rheumatoid arthritis. Interestingly, the barks of Liriodendron chinense (Hemsl.) Sarg was also found in folk to treat gout. However, further experimental studies remained to confirm its uric acid-lowering effects. AIM OF THE STUDY: The aim of the study was to evaluate the protective effect of ethanol extract of the barks of Liriodendron chinense (Hemsl.) Sarg (EELC) in a mouse model of hyperuricemic nephropathy (HN) and the involved mechanisms. MATERIALS AND METHODS: EELC at a respective dose of 250 mg/kg/d or 500 mg/kg/d were orally administered to HN mice induced by a mixture of adenine (160 mg/kg/d)/potassium oxonate (2.4 g/kg/d) for 21 days. At the end of the treatment, serum uric acid, kidney functions (serum creatinine, blood urea nitrogen and urine microalbumin), 24-h urine uric acid excretion, as well as kidney pathological changes were investigated by biochemical assay, histopathological score, immunofluorescence and histochemistry, RT-qPCR, and western blotting analysis. RESULTS AND DISCUSSION: Oral administration of EELC significantly lowered serum uric acid level at 500 mg/kg (185.75 ± 15.49 µmol/L of EELC vs. 238.28 ± 20.97 µmol/L of HN model, p < 0.01) in HN mice. EELC at 500 mg/kg also remarkably reduced the levels of serum creatinine (82.92 ± 7.86 µmol/L of EELC vs. 92.08 ± 6.13 µmol/L of HN model, p < 0.0001), blood urea nitrogen (21.50 ± 1.87 mmol/L of EELC vs. 29.40 ± 3.95 mmol/L of HN model, p < 0.001) and urine microalbumin (4.25 ± 0.40 mg/L of EELC vs. 5.95 ± 0.33 mg/L of HN model, p < 0.001) to improve renal function. It also attenuated renal fibrosis, especially the high-dose of EELC. Furthermore, EELC could inhibit the activation of NF-κB, ASK1/JNK/c-Jun, JAK2/STAT3 signaling pathways and reduce the release of pro-inflammatory cytokine TNF-α in the kidneys of HN mice. Additionally, EELC remarkably increased urine uric acid excretion of HN mice, which may be achieved by the upregulation of organic anion transporter 1 (OAT1), OAT3 and ATP-binding cassette subfamily G member 2 (ABCG2) proteins. CONCLUSIONS: EELC alleviated the progression of HN by suppressing the activation of NF-κB, ASK1/JNK/c-Jun and JAK2/STAT3 signaling pathway, reducing the infiltration of inflammatory factors and uric acid accumulation in the kidney.

Hydroalcoholic extract of Tagetes minuta L. inhibits inflammatory bowel disease through the activity of pheophytins on the NF-κB signalling pathway.

ETHNO-PHARMACOLOGICAL RELEVANCE: Species of the genus Tagetes are well known for their anti-inflammatory properties. Tagetes minuta "Huacatay" is an endemic species of South America that has been used in traditional medicine since ancient times as a remedy for stomach and intestinal discomfort. AIM OF THE STUDY: The aim of this study is to investigate the anti-inflammatory activity of the aqueous and hydroalcoholic extracts of the Huacatay, identifying the compounds responsible for this activity. MATERIALS AND METHODS: Anti-inflammatory activity of the compounds, fractions and extracts was evaluated in Hs 746T (stomach), HIEC-6 (intestine) and THP-1 (monocytes peripheral blood) cells by measuring their inhibitory capacity against the NF-κB production. RESULTS: Aqueous and hydroalcoholic extracts of Tagetes minuta displayed anti-inflammatory activity in vitro, the hydroalcoholic extract being the most active (IC50 between 59.72 and 66.42 µg/mL) in all cell lines. Bio-guided hydroalcoholic extract fractionation led to the isolation and characterisation of two pheophytins, pheophytin a (1) and 132-hydroxy pheophytin a (2). Both compounds inhibited the production of NF-κB with IC50 values in the low micromolar range, with an IC50 between 12.32 and 16.01 µM for compound 1 and 7.91-9.87 µM for compound 2. CONCLUSIONS: The two pheophytins isolated in this study inhibit the production of NF-κB, thus showing that the traditional anti-inflammatory use of Tagetes minuta can be proved through pharmacological assays. This contributes to understanding the anti-inflammatory activity of the Huacatay extracts and their use in the treatment of stomach and intestinal discomfort.

Sphaeranthus senegalensis DC: Evaluation of chemical constituents, oral safety, gastroprotective activity, and mechanism of action of its hydroethanolic extract.

ETHNOPHARMACOLOGICAL RELEVANCE: Sphaeranthus senegalensis DC is a seasonal herb with a spicy smell that grows wild in wet grounds of tropical Africa and Asia. The plant is used in folk medicine for the treatment of various diseases; that includes its use to treat gastric ulcers. AIM OF THE STUDY: This study aimed to investigate the chemical constituents of the hydroethanolic extract of Sphaeranthus senegalensis DC and evaluate its oral safety, gastroprotective activity, and mechanisms of action using laboratory models in rats and mice. MATERIALS AND METHODS: Hydroethanolic extract (70%) of the powdered whole dried material was prepared, and chemical constituents of the resultant extract (denoted HESs) standardized using the high-performance liquid chromatography (HPLC) method. The safety profile of HESs was assessed using 2000 mg/kg, oral (p.o.) for Hippocratic screening in mice, and 800 mg/kg, p.o. for 28 days subchronic toxicity assay in rats. The gastroprotective effect of HESs (25, 100, and 400 mg/kg, p.o.) was investigated using acidified ethanol, piroxicam, water immobilization stress, and acetic acid-induced ulcer models. The gastroprotective mechanisms of HESs were evaluated using its effect on gastric mucus protection, nitric oxide modulation, gastric juice secretory parameters, catalase and myeloperoxidase activities. Histological analysis of the stomach tissues was also carried out. RESULTS: The HPLC analysis indicated the presence of 25.94% phenolics (gallic acid, caffeic acid, and ferulic acid) and 14.53% flavonoids (rutin, morin, luteolin, quercetin, and apigenin). Hippocratic screening and the 28 days subchronic study indicated that HESs is generally safe. Result shows that oral administration of HESs (25, 100 and 400 mg/kg) alleviated the severity of the gastric ulcers induced by acidified ethanol by 35.65% (p < 0.05), 48.70% (p < 0.05) and 78.02% (p < 0.001) respectively; exhibited gastroprotective effect against the gastric lesions induced by piroxicam by 37.97% (p < 0.05), 53.27% (p < 0.05) and 76.23% (p < 0.001) respectively; and decreased the severity of the water immobilization stress-induced gastric ulcers by 32.43% (p < 0.05), 55.26% (p < 0.01) and 74.05% (p < 0.001) respectively, when compared to the vehicle control group. The mechanisms of action assays indicated that the gastroprotective activity was mediated mainly through gastroprotection, antisecretory, and antioxidant activities. Histological analysis showed it inhibited epithelial cell loss, vascular damage, and leucocyte infiltration. CONCLUSION: HESs contains useful phytochemicals, is safe, and exhibited significant gastroprotective action. The results provided justification for its claim in the treatment of gastric ulcers and its evaluation for potential application as a gastroprotective agent.

Anti-Vascular Inflammatory Effect of Ethanol Extract from Securinega suffruticosa in Human Umbilical Vein Endothelial Cells.

Securiniga suffruticosa is known as a drug that has the effect of improving the blood circulation and relaxing muscles and tendons, thereby protects and strengthen kidney and spleen. Therefore, in this study, treatment of Securiniga suffruticosa showed protective effect of inhibiting the vascular inflammation in human umbilical vein endothelial cells (HUVECs) by inducing nitric oxide (NO) production and endothelial nitric oxide synthase (eNOS) coupling pathway. In this study, Securiniga suffruticosa suppressed TNF-α (Tumor necrosis factor-α) induced protein and mRNA levels of cell adhesion molecules such as intracellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and Interleukin-6 (IL-6). Pretreatment of HUVEC with Securiniga suffruticosa decreased the adhesion of HL-60 cells to Ox-LDL (Oxidized Low-Density-Lipoprotein)-induced HUVEC. Moreover, Securiniga suffruticosa inhibited TNF-α induced intracellular reactive oxygen species (ROS) production. Securiniga suffruticosa also inhibited phosphorylation of IκB-α in cytoplasm and translocation of NF-κB (Nuclear factor-kappa B) p65 to the nucleus. Securiniga suffruticosa increased NO production, as well increased the phosphorylation of eNOS and Akt (protein kinase B) which are related with NO production. In addition, Securiniga suffruticosa increased the protein expression of GTPCH (Guanosine triphosphate cyclohydrolase Ⅰ) and the production of BH4 in HUVEC which are related with eNOS coupling pathway. In conclusion, Securiniga suffruticosa has a protective effect against vascular inflammation and can be a potential therapeutic agent for early atherosclerosis.

Ethanolic extract of Artemisia campestris subsp. glutinosa (Besser) Batt. inhibits HIV-1 replication in vitro through the activity of terpenes and flavonoids on viral entry and NF-κB pathway.

ETHNO-PHARMACOLOGICAL RELEVANCE: The genus Artemisia spp. is well known for its anti-infectious properties and its high content in anti-infectious compounds, like the well-known sweet wormwood (Artemisia annua L.). Another Artemisia species, Artemisia campestris subsp. glutinosa (Besser) Batt., field wormwood, has been traditionally used as medicinal plant in the Mediterranean region. AIM OF THE STUDY: The aim of this study is to investigate the anti-HIV activity of field wormwood, to identify the compounds responsible for this activity and their structure and mechanism of action. MATERIALS AND METHODS: Antiviral activity of isolated compounds and extracts was evaluated in HIV-1 infections of lymphoblastoid cells. We also evaluated the mechanism of action of isolated compounds. Viral entry was studied comparing the inhibitory effect of isolated compounds on wild type HIV-1 and VSV pseudotyped HIV-1. To assess the viral transcriptional effect, plasmids encoding luciferase reporter genes under the control of the whole genome of HIV-1 or NF-κB or Sp1 transcription factors were transfected in the presence of the compounds under evaluation. Finally, antioxidant activity was assessed by quantitation of reduced and total glutathione in treated cell cultures. RESULTS: Ethanolic and aqueous extracts of Artemisia campestris subsp. glutinosa (Besser) Batt. subsp. glutinosa displayed anti-HIV activity in vitro, although ethanolic extract was more powerful (IC50 14.62 µg/mL). Bio-guided ethanolic extract fractionation leads to the isolation and characterization of two terpenes, damsin and canrenone, and four flavonoids, 6, 2', 4'-trimethoxyflavone, acerosin, cardamonin and xanthomicrol. All the isolated compounds inhibited HIV-1 replication in vitro with IC50 values between the middle nanomolar and the low micromolar range. Their anti-HIV mechanism of action is due to the bloking of viral entry and/or transcription inhibition, without correlation with the antioxidant activity, through interference with the cellular transcription factors NF-κB and Sp1, which are targets that are not currently reached by antiretroviral therapy. CONCLUSION: We describe here the anti-HIV activity of field wormwood, Artemisia campestris subsp. glutinosa (Besser) Batt., and the isolation and study of the mechanism of action of two terpenes and four flavonoids, responsible, at least in part, for its activity, through the inhibition of two different cellular targets affecting the HIV replication cycle. The activity of these compounds in cellular targets could explain why plant extracts can be used in the treatment of different diseases. Besides, the presence of several compounds with dual and different mechanisms of action could prove useful in the treatment of HIV-1 infection, since it could aid to overcome drug resistances and simplify drug therapy. This work is a further step in understanding the anti-infectious activity of wormwood species and their use in treating infectious diseases.

Chemical composition, antioxidant, anti-inflammatory and antinociceptive activities of the ethanol extract of ripe fruits of Solanum lycocarpum St. Hil. (Solanaceae).

ETHNOPHARMACOLOGICAL RELEVANCE: Solanum lycocarpum St. Hil. (Solanaceae) is widely distributed in the Brazilian Cerrado and is used in folk medicine for treatment of inflammatory disorders, such as asthma and hepatitis, as weel as antirheumatic. AIM OF THE STUDY: The aims of this study were to evaluate the antioxidant, anti-inflammatory and antinociceptive activities of the ethanol extract (EE) obtained from the ripe fruits of S. lycocarpum and to identify its chemical constituents. MATERIALS AND METHODS: The extract was obtained by percolation with ethanol. This extract was analyzed by liquid chromatography coupled to a diode array detector and mass spectrometer (LC-DAD-MS) for identify its chemical constituents. The antioxidant activity was determined by the reaction with 1,1-diphenyl-2-picrylhydrazyl radical (DPPH). In vivo anti-inflammatory potential was assessed using carrageenan-induced paw edema model, while qualitative and quantitative histological analyses evaluated of the inflammatory infiltrate at different times and treatments. The antinociceptive effect of the EE was evaluated by acetic acid-induced abdominal writhing test, formalin-induced nociception and hot-plate test. RESULTS: The main compounds identified in EE were steroidal glycoalkaloids (such as robeneoside B or hydroxysolasonine isomers and solanandaine isomers), the aglycone alkaloids peiminine and solasodine, di- and tri-O-caffeoylquinic acid derivatives, O-coumaroyl caffeoylquinic acid derivatives, N1,N10-bis-(dihydrocaffeoyl)spermidine, di-O-hexoside, and hexonic acid. In addition, the EE showed significant antioxidant activity. Intraperitoneal (i.p.) treatment with EE (300 mg/kg) exhibited anti-inflammatory activity. Qualitative and quantitative histological analyses showed that EE significantly reduced the cell infiltrate in acute inflammation. The EE, in all doses evaluated, significantly reduced the abdominal contortions in mice. Besides, reduced licking time was found in both phases in the formalin test after treatment with EE (100 and 300 mg/kg). In addition, the opioid receptor antagonist naloxone reversed the antinociceptive activity of morphine in the both phases the test, but it did not reverse the antinociceptive activity of the EE. The EE (300 mg/kg) also caused an increase in the latency to response in the hot-plate test. CONCLUSION: The ripe fruits of S. lycocarpum exhibit antioxidant, anti-inflammatory, and antinociceptive activities, attributed mainly to the presence of alkaloids, such as solasodine and peiminine, as well as caffeoylquinic acids in their chemical composition. These results contribute to use of S. lycocarpum ripe fruits for the treatment of inflammatory and painful process.

Toxicity and antidiabetic activity of ethanolic extract of Sphagneticola trilobata (L.) Pruski flower in rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Plants are the source of medication for preventive, curative, protective or promotive purposes. Medicinal plants are an important source for generating of novel phytomedicine. They provide profound therapeutic benefits, more affordable treatments, effectiveness, less side effects and relatively low cost or less expensive and globally competitive. Using plant derived medicine is also relatively safe compared to synthetic medicines. Many plants have proved to successfully aid in the treatment of ailments including Sphagneticola triolobata (L.) Pruski. AIM OF STUDY: This study was therefore, designed to investigate acute and subacute toxicities, antidiabetic activity and also antioxidant activity of flower extract from S. triolobata (L.) Pruski. METHODS: This research investigates the toxicity and antidiabetic activity of Sphagnelicola trilobata (L.) Pruski flower ethanolic extract in rats. Acute toxicity was determined by a single oral administration of S. trilobata extract of 1500, 2000, and 2500 mg/kg body weight; and subacute toxicity by oral administration every two days for 14 days. Signs of toxicity and mortality were observed during 24 h and for 14 days. Hematological values and blood chemistry were also characterized. The antidiabetic activity was examined by orally administering S. trilobata extract of 250 mg/kg body weight to streptozotocin-induced diabetic rats on a daily basis for eight weeks; and the body weight, blood glucose, serum insulin, and lipid profiles were determined. The antioxidant activity of the extract was assessed by 1, 1-diphenyl-2-picryl-hydrazyl free radical scavenging assay. RESULTS AND CONCLUSION: The results demonstrated a median lethal dose (LD50) greater than 2500 mg/kg since there was no sign of toxicity and mortality in acute and subacute toxicity testing. The high LD50 indicated that S. trilobata flower ethanolic extract is safe for treatment of diabetes. There was no significant change in the body weight, hematological values, and blood chemistry of treated rats, compared with the control group. The diabetes-induced rats showed a significant reduction in blood glucose and triglyceride (p < 0.05). Meanwhile, the antioxidant activity of S. trilobata extract was lower than that of standard ascorbic acid.

Protective Effects of Facilitated Removal of Blood Alcohol and Acetaldehyde Against Liver Injury in Animal Models Fed Alcohol and Anti-HIV Drugs.

BACKGROUND: We previously developed enzyme nanoparticles (ENP) of alcohol metabolism. This study was to evaluate protective effects of facilitated removal of blood alcohol and/or acetaldehyde on anti-HIV drugs and alcohol-induced liver injuries. METHODS: ENP were prepared for degrading alcohol completely (ENP1) or partially into acetaldehyde (ENP2), which were applied to mice of acute binge or chronic-binge alcohol feeding in the presence of antivirals (ritonavir and lopinavir). Liver pathologies were examined to assess the protective effects of ENP. RESULTS: In the acute model, ENP1 and ENP2 reduced the blood alcohol concentration (BAC) by 41 and 32%, respectively, within 4 hr, whereas in control without ENP, BAC was reduced only by 15%. Blood acetaldehyde concentration (BADC) was increased by 39% in alcohol-fed mice treated with ENP2 comparing to control. No significant effects of the anti-HIV drugs on BAC or BADC were observed. Plasma alanine aminotransferase (ALT) and expression of liver TNF-α were both significantly increased in the alcohol-fed mice, which were normalized by ENP1. In the presence of the antivirals, ALT was partially reduced by ENP1 or ENP2. In the chronic model, inflammation, fatty liver, and ALT were increased, which were deteriorated by the antivirals. ENP1 partially reduced BAC, BADC, ALT, and expression of inflammation markers of TNF-α, F4/80, and IL-6 and lipogenic factors of ACC, LXRα, and SREBP1. ENP2 reduced BAC without significant effects on ALT, inflammation, or lipogenesis. Antivirals and alcohol synergistically increased expression of organelle stress markers of CHOP, sXBP-1, ATF6, and GCP60. ENP1 reduced BAC, CHOP, and sXbp-1. However, no effects of ENP1 were found on ATF6 or GCP60. CONCLUSIONS: Removal of blood alcohol and acetaldehyde by the ENP protects the liver against alcoholic injuries, and the protection is less effective in chronic alcohol and antiviral feeding due to additional drug-induced organelle stresses.

New Iridoid Derivatives from the Fruits of Cornus officinalis and Their Neuroprotective Activities.

Three previously undescribed iridoids, cornusfurals A⁻C, were isolated from the ethanolic extracts of fruits of Cornus officinalis. Their structures were elucidated by spectroscopic methods, including one-dimensional and two-dimensional nuclear magnetic resonance, ultraviolet spectroscopy, infrared spectroscopy, and mass spectrometry. The neuroprotective activity was evaluated by measuring corticosterone-induced damage in PC12 cells. The results showed that cornusfural B decreased corticosterone-induced PC12 cell damage compared with that in model cells.

Development and Validation of a HPLC-UV Method for Extraction Optimization and Biological Evaluation of Hot-Water and Ethanolic Extracts of Dendropanax morbifera Leaves.

Dendropanax morbifera Leveille (Araliaceae) has been used in traditional oriental remedies for cancer, inflammation, diabetes, and thrombosis. However, a validated analytical method, standardization, and optimization of extraction conditions with respect to biological activity have not been reported. In this study, a simple and validated HPLC method for identifying and quantifying active substances in D. morbifera was developed. Hot water and ethanolic D. morbifera leaf extracts from different production regions were prepared and evaluated with regard to their chemical compositions and biological activities. The contents of active compounds such as rutin and chlorogenic acid were determined in four samples collected from different regions. The 80% ethanolic extract showed the best antioxidant activity, phenolic content, reducing power, and xanthine oxidase (XO) inhibitory activity. The validated HPLC method confirmed the presence of chlorogenic acid and rutin in D. morbifera leaf extracts. The antioxidant and XO inhibitory activity of D. morbifera extract could be attributed to the marker compounds. Collectively, these results suggest that D. morbifera leaves could be beneficial for the treatment or prevention of hyperuricemia-related disease, and the validated HPLC method could be a useful tool for the quality control of food or drug formulations containing D. morbifera.

[Analyze nanofiltration separation rule of chlorogenic acid from low concentration ethanol by Donnan effect and solution-diffusion effect].

To separate chlorogenic acid from low concentration ethanol and explore the influence of Donnan effect and solution-diffusion effect on the nanofiltration separation rule. The experiment showed that solution pH and ethanol volume percent had influences on the separation of chlorogenic acid. Within the pH values from 3 to 7 for chlorogenic acid in 30% ethanol, the rejection rate of chlorogenic acid was changed by 70.27%. Through the response surface method for quadratic regression model, an interaction had been found in molecule weight cut-off, pH and ethanol volume percent. In fixed nanofiltration apparatus, the existence states of chlorogenic acid determinedits separation rules. With the increase of ethanol concentration, the free form chlorogenic acid was easily adsorbed, dissolved on membrane surface and then caused high transmittance due to the solution-diffusion effect. However, at the same time, due to the double effects of Donnan effect and solution-diffusion effect, the ionic state of chlorogenic acid was hard to be adsorbed in membrane surface and thus caused high rejection rate. The combination of Box-Behnken design and response surface analysis can well optimize the concentrate process by nanofiltration, and the results showed that nanofiltration had several big advantages over the traditional vacuum concentrate technology, meanwhile, and solved the problems of low efficiency and serious component lossesin the Chinese medicines separation process for low concentration organic solvent-water solution.

A dynamic metabolic flux analysis of ABE (acetone-butanol-ethanol) fermentation by Clostridium acetobutylicum ATCC 824, with riboflavin as a by-product.

The present study reveals that supplementing sodium acetate (NaAc) strongly stimulates riboflavin production in acetone-butanol-ethanol (ABE) fermentation by Clostridium acetobutylicum ATCC 824 with xylose as carbon source. Riboflavin production increased from undetectable concentrations to ∼0.2 g L-1 (0.53 mM) when supplementing 60 mM NaAc. Of interest, solvents production and biomass yield were also promoted with fivefold acetone, 2.6-fold butanol, and 2.4-fold biomass adding NaAc. A kinetic metabolic model, developed to simulate ABE biosystem, with riboflavin production, revealed from a dynamic metabolic flux analysis (dMFA) simultaneous increase of riboflavin (ribA) and GTP (precursor of riboflavin) (PurM) synthesis flux rates under NaAc supplementation. The model includes 23 fluxes, 24 metabolites, and 72 kinetic parameters. It also suggested that NaAc condition has first stimulated the accumulation of intracellular metabolite intermediates during the acidogenic phase, which have then fed the solventogenic phase leading to increased ABE production. In addition, NaAc resulted in higher intracellular levels of NADH during the whole culture. Moreover, lower GTP-to-adenosine phosphates (ATP, ADP, AMP) ratio under NaAc supplemented condition suggests that GTP may have a minor role in the cell energetic metabolism compared to its contribution to riboflavin synthesis.

Optimization of enzymatic hydrolysis and fermentation conditions for improved bioethanol production from potato peel residues.

The aim of this work was the optimization of the enzyme hydrolysis of potato peel residues (PPR) for bioethanol production. The process included a pretreatment step followed by an enzyme hydrolysis using crude enzyme system composed of cellulase, amylase and hemicellulase, produced by a mixed culture of Aspergillus niger and Trichoderma reesei. Hydrothermal, alkali and acid pretreatments were considered with regards to the enhancement of enzyme hydrolysis of potato peel residues. The obtained results showed that hydrothermal pretreatment lead to a higher enzyme hydrolysis yield compared to both acid and alkali pretreatments. Enzyme hydrolysis was also optimized for parameters such as temperature, pH, substrate loading and surfactant loading using a response surface methodology. Under optimized conditions, 77 g L-1 of reducing sugars were obtained. Yeast fermentation of the released reducing sugars led to an ethanol titer of 30 g L-1 after supplementation of the culture medium with ammonium sulfate. Moreover, a comparative study between acid and enzyme hydrolysis of potato peel residues was investigated. Results showed that enzyme hydrolysis offers higher yield of bioethanol production than acid hydrolysis. These results highlight the potential of second generation bioethanol production from potato peel residues treated with onsite produced hydrolytic enzymes. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:397-406, 2017.

Hybrid SSF/SHF Processing of SO2 Pretreated Wheat Straw-Tuning Co-fermentation by Yeast Inoculum Size and Hydrolysis Time.

Wheat straw is one of the main agricultural residues of interest for bioethanol production. This work examines conversion of steam-pretreated wheat straw (using SO2 as a catalyst) in a hybrid process consisting of a short enzymatic prehydrolysis step and a subsequent simultaneous saccharification and fermentation (SSF) step with a xylose-fermenting strain of Saccharomyces cerevisiae. A successful process requires a balanced design of reaction time and temperature in the prehydrolysis step and yeast inoculum size and temperature in the SSF step. The pretreated material obtained after steam pretreatment at 210 °C for 5 min using 2.5 % SO2 (based on moisture content) showed a very good enzymatic digestibility at 45 °C but clearly lower at 30 °C. Furthermore, the pretreatment liquid was found to be rather inhibitory to the yeast, partly due to a furfural content of more than 3 g/L. The effect of varying the yeast inoculum size in this medium was assessed, and at a yeast inoculum size of 4 g/L, a complete conversion of glucose and a 90 % conversion of xylose were obtained within 50 h. An ethanol yield (based on the glucan and xylan in the pretreated material) of 0.39 g/g was achieved for a process with this yeast inoculum size in a hybrid process (10 % water-insoluble solid (WIS)) with 4 h prehydrolysis time and a total process time of 96 h. The obtained xylose conversion was 95 %. A longer prehydrolysis time or a lower yeast inoculum size resulted in incomplete xylose conversion.

Simultaneous Saccharification and Fermentation of Sugar Beet Pulp for Efficient Bioethanol Production.

Sugar beet pulp, a byproduct of sugar beet processing, can be used as a feedstock in second-generation ethanol production. The objective of this study was to investigate the effects of pretreatment, of the dosage of cellulase and hemicellulase enzyme preparations used, and of aeration on the release of fermentable sugars and ethanol yield during simultaneous saccharification and fermentation (SSF) of sugar beet pulp-based worts. Pressure-thermal pretreatment was applied to sugar beet pulp suspended in 2% w/w sulphuric acid solution at a ratio providing 12% dry matter. Enzymatic hydrolysis was conducted using Viscozyme and Ultraflo Max (Novozymes) enzyme preparations (0.015-0.02 mL/g dry matter). Two yeast strains were used for fermentation: Ethanol Red (S. cerevisiae) (1 g/L) and Pichia stipitis (0.5 g/L), applied sequentially. The results show that efficient simultaneous saccharification and fermentation of sugar beet pulp was achieved. A 6 h interval for enzymatic activation between the application of enzyme preparations and inoculation with Ethanol Red further improved the fermentation performance, with the highest ethanol concentration reaching 26.9 ± 1.2 g/L and 86.5 ± 2.1% fermentation efficiency relative to the theoretical yield.

Beneficial Effects of Trillium govanianum Rhizomes in Pain and Inflammation.

Trillium govanianum rhizome is used as an analgesic and anti-inflammatory remedy in traditional medicine in northern Pakistan. In an attempt to establish its medicinal value, the present research evaluated the analgesic and anti-inflammatory potential of T. govanianum. The in vivo anti-inflammatory activity of extract and fractions was investigated in the carrageenan induced paw edema assay. The in vitro suppression of oxidative burst of extract, fractions and isolated compounds was assessed through luminol-enhanced chemiluminescence assay. The in vivo analgesic activity was assayed in chemical and thermal induced nociceptive pain models. The crude methanol extract and its solvent fractions showed anti-inflammatory and analgesic responses, exhibited by significant amelioration of paw edema and relieve of the tonic visceral chemical and acute phasic thermal nociception. In the oxidative burst assay, based on IC50, the crude methanol extract and n-butanol soluble fraction produced a significant inhibition, followed by chloroform and hexane soluble fractions as compared to ibuprofen. Similarly, the isolated compounds pennogenin and borassoside E exhibited significant level of oxidative burst suppressive activity. The in vivo anti-inflammatory and analgesic activities as well as the in vitro inhibition of oxidative burst validated the traditional use of T. govanianum rhizomes as a phytotherapeutic remedy for both inflammatory conditions and pain. The observed activities might be attributed to the presence of steroids and steroid-based compounds. Therefore, the rhizomes of this plant species could serve as potential novel source of compounds effective for alleviating pain and inflammation.

ISOLATION AND CHARACTERIZATION OF CHEMICAL CONSTITUENTS FROM CHRYSOPHYLLUM ALBIDUM G. DON-HOLL. STEM-BARK EXTRACTS AND THEIR ANTIOXIDANT AND ANTIBACTERIAL PROPERTIES.

BACKGROUND: The plant, Chrysophyllum albidum is indigenous to Nigeria and its stem-bark has wide application in traditional medicine for the treatment of infections and oxidative stress related diseases. The aim of the study was to isolate the chemical constituents responsible for the antioxidant and antibacterial activity from the stem-bark of the plant in order to justify some of its ethnomedicinal uses. MATERIALS AND METHODS: Crude extract of stem-bark of Chrysophyllum albidum obtained from 80% ethanol was successively partitioned with ethyl acetate (EtOAc) and n-butanol. The solvent fractions and isolated compounds were tested for antioxidant property using 2-2-diphenyl-1-picrylhydrazyl. Antibacterial activities were also assessed by means of agar-diffusion and broth micro dilution methods. EtOAc fraction was repeatedly fractionated on column chromatography to afford four compounds and their chemical structures were established using NMR (1D and 2D) and MS. RESULTS: Chromatographic fractionation of EtOAc fraction with the highest antioxidant and antimicrobial activities afforded stigmasterol (1),: epicatechin (2),: epigallocatechin (3): and procyanidin B5 (4).: Procyanidin B5 isolated for the first time from genus Chrysophyllum demonstrated the highest antioxidant activity with IC50 values of 8.8 µM and 11.20 µM in DPPH and nitric oxide assays respectively and equally demonstrated the highest inhibitory activity against Escherichia coli (MIC 156.25 µg/mL), Staphylococcus aureus (MIC 156.25 µg/mL), Pseudomonas aeruginosa (MIC 625 µg/mL) and Bacillus subtilis (MIC 156.25 µg/mL). CONCLUSION: The antibacterial and antioxidant activities of epicatechin, epigallocatechin and procyanidin B5 isolated from Chrysophyllum albidum stem-bark validate the folkloric uses.

Clinacanthus nutans (Burm. f.) Lindau Ethanol Extract Inhibits Hepatoma in Mice through Upregulation of the Immune Response.

Clinacanthans nutans (Burm. f.) Lindau is a popular medicinal vegetable in Southern Asia, and its extracts have displayed significant anti-proliferative effects on cancer cells in vitro. However, the underlying mechanism for this effect has yet to be established. This study investigated the antitumor and immunomodulatory activity of C. nutans (Burm. f.) Lindau 30% ethanol extract (CN30) in vivo. CN30 was prepared and its main components were identified using high-performance liquid chromatography (HPLC) and mass spectrometry (LC/MS/MS). CN30 had a significant inhibitory effect on tumor volume and weight. Hematoxylin and eosin (H & E) staining and TUNEL assay revealed that hepatoma cells underwent significant apoptosis with CN30 treatment, while expression levels of proliferation markers PCNA and p-AKT were significantly decreased when treated with low or high doses of CN30 treatment. Western blot analysis of PAPR, caspase-3, BAX, and Bcl2 also showed that CN30 induced apoptosis in hepatoma cells. Furthermore, intracellular staining analysis showed that CN30 treatment increased the number of IFN-γ⁺ T cells and decreased the number of IL-4⁺ T cells. Serum IFN-γ and interleukin-2 levels also significantly improved. Our findings indicated that CN30 demonstrated antitumor properties by up-regulating the immune response, and warrants further evaluation as a potential therapeutic agent for the treatment and prevention of cancers.

Antileishmanial activity of compounds produced by endophytic fungi derived from medicinal plant Vernonia polyanthes and their potential as source of bioactive substances.

The purpose of this work was to evaluate the antileishmanial activity of endophytic fungi isolated from leaves of Vernonia polyanthes plant and their prospective use in the discovery of bioactive compounds. Sixteen endophytes were isolated by using potato dextrose agar medium and submitted to cultivation in rice medium. The fungal cultures were extracted with ethanol and used as crude extracts for testing their antileishmanial activity. The most active ethanol extract was obtained from P2-F3 strain, which was identified as Cochliobolus sativus by ITS rRNA gene sequence data. Followed by a bioassay-guided fractionation, the cochlioquinone A, isocochlioquinone A and anhydrocochlioquinone A compounds were isolated from the crude extracts and demonstrated to inhibit the parasites. From the present work, it is possible to conclude that endophytic fungi derived from medicinal plant V. polyanthes may be considered promising source for the discovery of bioactive compounds.