Isolation, Purification, Fractionation, and Hepatoprotective Activity of Polygonatum Polysaccharides.

In this study, three homogeneous fractions, PSP-N-b-1, PSP-N-b-2, and PSP-N-c-1, were obtained from an aqueous extract of Polygonatum using DEAE cellulose column chromatography, CL-6B agarose gel chromatography, and Sephadex G100 chromatography. Their monosaccharide compositions and molecular weights were analyzed. The results revealed that PSP-N-b-1, PSP-N-b-2, and PSP-N-c-1 are primarily composed of six monosaccharides: Man (mannose), GlcA (glucuronic acid), Rha (rhamnose), GalA (galacturonic acid), Glc (glucose), and Ara (arabinose), with molecular weights of 6.3 KDa, 5.78 KDa, and 3.45 KDa, respectively. Furthermore, we observed that Polygonatum polysaccharides exhibited protective effects against CCL4-induced liver damage in HepG2 cells in vitro, operating through both anti-oxidant and anti-inflammatory mechanisms. Our research findings suggest that Polygonatum polysaccharides may emerge as a promising option in the development of hepatoprotective drugs or functional foods with anti-inflammatory and antioxidant properties.

Isolation, purification, and physicochemical characterization of Polygonatum polysaccharide and its protective effect against CCl4-induced liver injury via Nrf2 and NF-κB signaling pathways.

This study aimed to provide scientific evidence that Polygonatum polysaccharide can be developed as a dietary supplement and medication for treating liver injuries. A water-soluble polysaccharide (PSP-N-c-1), with an average molecular weight of 3.45 kDa, was isolated and purified from the water extract of Polygonatum using DEAE cellulose column chromatography, CL-6B agarose gel chromatography, and Sephadex G100 chromatography. High-performance liquid chromatography, gas chromatography-mass spectrometry, and nuclear magnetic resonance spectroscopy analyses revealed that PSP-N-c-1 might be linear α-(1 â†’ 4)-glucans with α-Glcp residues linked to the backbone at C-6. In vitro experiments revealed that PSP-N-c-1 exhibited protective effects against CCl4-induced damage in HepG2 cells. In vivo experiments demonstrated that PSP-N-c-1 exhibited a hepatoprotective effect by enhancing antioxidant enzyme activity, inhibiting lipid peroxidation, and reducing the activity of pro-inflammatory mediators. Besides, PSP-N-c-1 could attenuate oxidative stress and inflammatory responses by activating the Nrf2-mediated signaling pathways and regulating the TLR4-mediated NF-κB signaling pathways. These findings demonstrated that PSP-N-c-1 may serve as a supplement for alleviating chemical liver damage.

Effect of Temperature, pH, and aw on Cereulide Synthesis and Regulator Genes Transcription with Respect to Bacillus cereus Growth and Cereulide Production.

Bacillus cereus is a food-borne pathogen that can produce cereulide in the growth period, which causes food poisoning symptoms. Due to its resistance to heat, extreme pH, and proteolytic enzymes, cereulide poses a serious threat to food safety. Temperature, pH, and aw can influence cereulide production, but there is still a lack of research with multi-environmental impacts. In this study, the effects of temperature (15~45 °C), pH (5~8), and aw (0.945~0.996) on the emetic reference strain B. cereus F4810/72 growth, cereulide production, relevant ces genes (cesA, cesB, cesP), and transcription regulators genes (codY and abrB) expression at transcription level were studied. B. cereus survived for 4~53 h or grew to 6.85~8.15 log10 CFU/mL in environmental combinations. Cereulide accumulation was higher in mid-temperature, acidic, or high aw environments. Increased temperature resulted in a lower cereulide concentration at pH 8 or aw of 0.970. The lowest cereulide concentration was found at pH 6.5 with an increased aw from 0.970 to 0.996. Water activity had a strong effect on transcriptional regulator genes as well as the cesB gene, and temperature was the main effect factor of cesP gene expression. Moreover, environmental factors also impact cereulide synthesis at transcriptional levels thereby altering the cereulide concentrations. The interaction of environmental factors may result in the survival of B. cereus without growth for a period. Gene expression is affected by environmental factors, and temperature and pH may be the main factors influencing the correlation between B. cereus growth and cereulide formation. This study contributed to an initial understanding of the intrinsic link between the impact of environmental factors and cereulide formation and provided valuable information for clarifying the mechanism of cereulide synthesis in combined environmental conditions.

Exposure of Salmonella enterica serovar 1,4,[5],12:i:- to benzalkonium chloride leads to acquired resistance to this disinfectant and antibiotics.

AIMS: Disinfectants such as benzalkonium chloride (BC), extensively used in animal farms and food-processing industries, contribute to the development of adaptive and cross-resistance in foodborne pathogens, posing a serious threat to food safety and human health. The purpose of this study is to explore whether continuous exposure of Salmonella enterica serovar 1,4,[5],12:i:- (S. 1,4,[5],12:i:-) to sublethal concentrations of BC could result in acquired resistance to this agent and other environmental stresses (e.g. antibiotics, heat, and acid). METHODS AND RESULTS: BC tolerance increased in all tested strains after exposure to gradually increasing concentrations of BC, with increases in minimum inhibitory concentrations between two and sixfold. The survival rate of BC-adapted strains was significantly (P < 0.05) higher than that of their wild-type (non-adapted) counterparts in lethal concentrations of BC. In addition, significant reductions (P < 0.05) in zeta potential were observed in BC-adapted strains compared to wild-type ones, indicating that a reduction in cell surface charge was a cause of adaptative resistance. More importantly, two BC-adapted strains exhibited increased antibiotic resistance to levofloxacin, ceftazidime, and tigecycline, while gene mutations (gyrA, parC) and antibiotic efflux-related genes (acrB, mdsA, mdsB) were detected by genomic sequencing analysis. Moreover, the tolerance of BC-adapted strains to heat (50, 55, and 60°C) and acid (pH 2.0, 2.5) was strain-dependent and condition-dependent. CONCLUSIONS: Repeated exposure to sublethal concentrations of BC could result in the emergence of BC- and antibiotic-resistant S. 1,4,[5],12:i:- strains.

Crypt-like patterned electrospun nanofibrous membrane and probiotics promote intestinal epithelium models close to tissues.

In vitro intestinal epithelium models have drawn great attention to investigating intestinal biology in recent years. However, the difficulty to maintain the normal physiological status of primary intestinal epithelium in vitro limits the applications. Here, we designed patterned electrospun polylactic acid (PLA) nanofibrous membranes with crypt-like topography and mimic ECM fibrous network to support crypt culture and construct in vitro intestinal epithelium models. The patterned electrospun PLA nanofibrous membranes modified with Matrigels at 0 °C showed high biocompatibility and promoted cell growth and proliferation. The constructed duodenum epithelium models and colon epithelium models on the patterned electrospun PLA nanofibrous membranes expressed the typical differentiation markers of intestinal epithelia and the gene expression levels were close to the original tissues, especially with the help of probiotics. The constructed intestinal epithelium models could be used to assess probiotic adhesion and colonization, which were verified to show significant differences with the Caco-2 cell models due to the different cell types. These findings provide new insights and a better understanding of the roles of biophysical, biochemical, and biological signals in the construction of in vitro intestinal epithelium models as well as the potential applications of these models in the study of host-gut microbes interactions. KEY POINTS: • Patterned electrospun scaffold has crypt-like topography and ECM nanofibrous network. • Matrigels at 0°C modify scaffolds more effectively than at 37°C. • Synergy of biomimic scaffold and probiotics makes in vitro model close to tissue.

Assay development considerations to improve drug tolerance in direct competitive ligand binding neutralizing antibody assays, pretreatment strategies.

Neutralizing anti-drug antibodies (ADAs) may affect safety, efficacy, and pharmacokinetic profile of a biotherapeutic drug and thus their assessment is of particular importance during immunogenicity testing. Neutralizing antibody (NAb) assays typically rely on NAbs ability to block the drug-target interaction. Higher NAb concentration and/or higher binding affinity of NAb to the drug, lowers the drug-target binding interaction. However, in the presence of high concentrations of residual circulating drug, as often seen for drugs with longer half-lives or in repeat-dose studies, NAbs may exist as drug bound complexes. In direct NAb assay formats, the NAb-drug complexes present in the sample could result in the NAb being unable to block the drug-target interaction eventually leading to a false negative response. The residual free circulating drug present in the sample may bind to the target in the NAb assay thereby competing with the drug used in the assay and inhibiting the assay signal, leading to a false positive response. For traditional ADA assays, multiple approaches involving acid treatment have been described to mitigate circulating drug interference issue. Here, we report two acid-treatment approaches that utilize the Dynabeads extraction with acid dissociation and Affinity Capture Elution (ACE) principle to improve drug tolerance in NAb assays.

Different cellular fatty acid pattern and gene expression of planktonic and biofilm state Listeria monocytogenes under nutritional stress.

Listeria monocytogenes is a Gram-positive bacterium frequently involved in food-borne disease outbreaks and is widely distributed in the food-processing environment. This work aims to depict the impact of nutrition deficiency on the survival strategy of L. monocytogenes both in planktonic and biofilm states. In the present study, cell characteristics (autoaggression, hydrophobicity and motility), membrane fatty acid composition of MRL300083 (Lm83) in the forms of planktonic and biofilm-associated cells cultured in TSB-YE and 10-fold dilutions of TSB-YE (DTSB-YE) were investigated. Additionally, the relative expression of related genes were also determined by RT-qPCR. It was observed that cell growth in different bacterial life modes under nutritional stress rendered the cells a distinct phenotype. The higher autoaggression (AAG) and motility of the planktonic cells in DTSB-YE is associated with better biofilm formation. An increased proportion of unsaturated fatty acid/saturated fatty acid (USFA/SFA) indicates more fluidic biophysical properties for cell membranes of L. monocytogenes in planktonic and biofilm cells in DTSB-YE. Biofilm cells produced a higher percentage of USFA and straight fatty acids than the corresponding planktonic cells. An appropriate degree of membrane fluidity is crucial for survival, and alteration of membrane lipids is an essential adaptive response. The adaptation of bacteria to stress is a multifactorial cellular process, the expression of flagella-related genes fliG, fliP, flgE and the two-component chemotactic system cheA/Y genes of planktonic cells in DTSB-YE significantly increased compared to that in TSB-YE (p < 0.05). This study provides new information on the role of the physiological adaptation and gene expression of L. monocytogenes for planktonic and biofilm growth under nutritional stress.

Heat shock in Cronobacter sakazakii induces direct protection and cross-protection against simulated gastric fluid stress.

The bacterial heat shock response in foodborne pathogens is caused by exposure to higher temperatures which poses a great threat to food safety because it can undermine food processing interventions and host defense. The study assessed the heat and acid resistance of Cronobacter sakazakii following heat shock (53 °C for 15min). Inactivation curves of the heat-shocked and non-shocked C. sakazakii cells at four temperatures (56, 58, 60, and 62 °C) and simulated gastric fluid (SGF, pH 3.0) were examined and fitted with the log-linear model and the Weibull model. The inactivation parameters obtained on the basis of the Weibull model showed that heat shock significantly (p < 0.05) increased the values of δ (time to reach 1 log reduction) and t3d (time to reach 3 log reduction) under thermal and acid inactivation. The results proved that heat shock provided C. sakazakii direct protection from a more adverse heat challenge and cross-protection from SGF, i.e. there was a heat shock response. Results of sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) revealed that seven protein species showed enhanced expression, while four protein species showed decreased expression in the heat-shocked compared to the non-heat-shocked C. sakazakii cells. Quantitative real-time reverse transcriptase PCR (RT-qPCR) revealed upregulation of six stress related genes, ibpA, ibpB (both encoding molecular chaperons), Hsp15 (encoding heat shock protein), Hsp20 (encoding small heat-shock protein), HspQ (encoding proteases) and rpoS (encoding stationary phase sigma factor), following heat shock treatment. In addition, heat shock induced an increase proportion of saturated fatty acids (SFA), cyclic fatty acids (CFA) and the ratio of saturated fatty acids to unsaturated fatty acids (SFA/USFA), whereas reducing the proportion of unsaturated fatty acids (USFA). Consequently, establishment of inactivation models of C. sakazakii could provide data support for quantitative microbial risk assessment (QMRA). Exploration of enhanced resistance mechanisms might provide clues for prevention and control of contamination by heat-shocked C. sakazakii.

Detection and Genotyping of Francisella tularensis in Animal Hosts and Vectors from Six Different Natural Landscape Areas, Gansu Province, China.

OBJECTIVE: Tularemia, also known as hare fever, is caused by the bacterium Francisella tularensis (F. tularensis) transmitted through diseased wild animals, blood sucking insects, or contaminated water or food, which is distributed worldwide. The purpose of this study was to investigate F. tularensis infection in animal hosts and vectors from six different natural landscape areas in Gansu Province and to identify the genotypes of the detected F. tularensis. METHODS: Rodents were captured by snap traps, and ticks were collected by dragging a cloth over the vegetation or from domestic animals. After species identification, DNA was isolated from the captured animals and detected by nested PCR assays targeting the F. tularensis fopA gene. The positive samples were further amplified to discriminate the species, and another two short-sequence tandem repeat regions (SSTR) were amplified to identify their genotypes. All positive fragments were sequenced and analyzed by ClustalX (5.0) and DNAClub software. RESULTS: A total of 407 rodents of 12 species were captured, among which six rodent species were positive for F. tularensis, with an overall prevalence of 3.93%. The geographical difference in infection rate was statistically significant. At the SSTR9 locus, there were 7 genotypes among positive rodent samples. A total of 1864 ticks were tested for evidence of tularemia by nested PCR assays, 69 of which were positive, with an average positive rate of 3.70% for F. tularensis in ticks. The positive rates were significantly different among different regions. Seven genotypes were identified at the SSTR9 locus, one of which seemed dominant in positive tick samples. All positive samples had the same genotype at the SSTR16 locus. CONCLUSION: There is natural infection of F. tularensis among animal vectors and hosts in Gansu Province, with diverse genotypes.

Epimedium flavonoids protect neurons and synapses in the brain via activating NRG1/ErbB4 and BDNF/Fyn signaling pathways in a chronic cerebral hypoperfusion rat model.

Cerebral hypoperfusion is a common feature of cerebral small vascular disease (CSVD), which has been considered as one of the causes of cognitive decline in recent years. Epimedium flavonoids (EF) are the main ingredients extracted from Epimedium. The purpose of this study was to investigate the effects of EF on cognitive impairment, and the underlying mechanisms in rats with permanent occlusion of the bilateral common carotid artery (2VO). EF (50, 100, and 200 mg/kg) was intragastrically administered for 12 weeks starting 2 weeks after 2VO surgery. The results showed that EF treatment improved learning and memory impairment in 2VO rats evaluated by novel object recognition and Y-maze tests. NeuN immunohistochemical staining indicated that EF alleviated neuronal loss in the hippocampus and cerebral cortex of 2VO rats. MAP-2 immunofluorescence staining and western blotting showed that EF protected neuronal dendrites and increased the expression of cytoskeleton proteins MAP-2 and NF200 in the hippocampus of 2VO rats. Moreover, EF protected the synapse ultrastructure detected by transmission electron microscopy, and increased the expression of synaptic plasticity-related proteins, including synaptophysin, synaptotagmin-I, synapsin I, PSD-95, p-NMDA2B, and p-CaMKII-α in the hippocampus of 2VO rats. In addition, EF increased the expression of neuregulin-1 (NRG-1), p-ErbB4, brain-derived neurotrophic factor (BDNF), p-Fyn, PI3K, p-Akt, and p-CREB in the hippocampus of 2VO rats. These results suggest that EF may protect neurons and synapses by activating the NRG1/ErbB4, BDNF/Fyn, and P13 K/Akt/CREB pathways in the hippocampus and cerebral cortex, thus improving cognitive impairment induced by chronic cerebral hypoperfusion. EF may be a potential candidate drug for chronic cerebral hypoperfusion and CSVD therapy.

Epimedium flavonoids improve cognitive impairment and white matter lesions induced by chronic cerebral hypoperfusion through inhibiting the Lingo-1/Fyn/ROCK pathway and activating the BDNF/NRG1/PI3K pathway in rats.

Chronic cerebral hypoperfusion is a common cause of cerebral small vascular disease (CSVD). White matter (WM) lesions are the typical pathological manifestation of CSVD and contribute to cognitive decline. Epimedium flavonoids (EF) are the main component in Epimedium brevicornu Maxim., which is commonly used in traditional Chinese medicine. The purpose of this study was to investigate the effects of EF on cognitive impairment and the underlying mechanisms in a CSVD rat model induced with chronic cerebral hypoperfusion. The model was established by permanent bilateral common carotid artery occlusion (2VO) in rats. EF (50, 100, and 200 mg/kg) was intragastrically administered once a day for 12 weeks starting 2 weeks after 2VO surgery. The learning and memory capacity of the rats were measured using the Morris water maze and step-through tests. WM lesions were observed by MRI-diffusion tensor imaging, transmission electron microscopy, and LFB staining. Oligodendrocytes were detected by immunohistochemistry. Western blotting assay was used to determine the level of protein expression. The results showed that EF significantly improved learning and memory impairment, alleviated WM nerve fiber injuries and demyelination, and increased the number of mature oligodendrocytes in the corpus callosum, subcortical WM, and periventricular WM in 2VO rats. Mechanistically, EF reduced the expression of Lingo-1 and ROCK2 and increased the levels of phosphorylated (p-) Fyn, brain-derived neurotrophic factor (BDNF), TrkB, neuregulin-1 (NRG-1), p-ErbB4, PI3K p85 and p110α, p-Akt, and p-CREB in the corpus callosum of 2VO rats. These results suggest that EF may improve cognitive impairment and WM lesions induced by chronic cerebral hypoperfusion through inhibiting the Lingo-1/Fyn/ROCK pathway and activating the BDNF/TrkB, NRG-1/ErbB4, and the downstream PI3K/Akt/CREB pathways in WM. Thus, EF can be used as a potential neuroprotective agent in CSVD therapy.

Gambogenic acid exerts anticancer effects in cisplatin­resistant non­small cell lung cancer cells.

Non­small cell lung cancer (NSCLC) is the most common type of lung cancer and the most common cause of mortality in patients with lung cancer. The efficacy of cisplatin­based chemotherapy in NSCLC is limited by drug resistance, therefore, the development of novel anticancer agents is required to overcome cisplatin resistance. The present study investigated the anticancer activity of gambogenic acid (GNA), derived from gamboge, in the cisplatin­resistant NSCLC cell line A549/Cis. GNA was revealed to have a potent inhibitory effect on cell growth in A549/Cis cells by blocking the cell cycle and inducing apoptosis. The investigation of the molecular mechanisms identified that GNA arrested the cell cycle at the G1 phase through the downregulation of cyclin Ds, cyclin dependent kinase (CDK)4 and CDK6, and the upregulation of p53 and p21. In addition, GNA induced apoptosis by increasing the activation of caspase 3 and caspase 7, in addition to the cleavage of poly(ADP­ribose) polymerase. The results of the present study supported the potential application of GNA in cisplatin­resistant NSCLC.

Cornel Iridoid Glycoside Protects Against White Matter Lesions Induced by Cerebral Ischemia in Rats via Activation of the Brain-Derived Neurotrophic Factor/Neuregulin-1 Pathway.

BACKGROUND: Ischemic stroke often induces profound white matter lesions, resulting in poor neurological outcomes and impaired post-stroke recovery. The present study aimed to investigate the effects of cornel iridoid glycoside (CIG), a major active component extracted from Cornus officinalis, on the white matter injury induced by ischemic stroke and further investigate its neuroprotective mechanisms. METHODS: Adult male Sprague-Dawley rats underwent middle cerebral artery occlusion (MCAO) surgery for 2 h, followed by reperfusion. Rats were intragastrically administered CIG (60 mg/kg and 120 mg/kg) beginning 6 h afters reperfusion, once daily for seven days. A series of behavioral tests (modified neurological severity scores test, object recognition test, adhesive removal test, and beam walking test) were performed to evaluate the neurological functioning in MCAO rats. Histology of the white matter was studied using luxol fast blue staining and transmission electron microscopy. Immunohistochemical staining was performed to assess myelin loss, oligodendrocyte maturation, and glial activation. Activation of the brain-derived neurotrophic factor (BDNF)/neuregulin-1 (NRG1) pathway was evaluated by Western blotting. RESULTS: CIG treatment remarkably decreased the neurological deficit score, accelerated the recovery of somatosensory and motor functions, and ameliorated the memory deficit in MCAO rats. Furthermore, CIG alleviated white matter lesions and demyelination, increased myelin basic protein expression and the number of mature oligodendrocytes, and decreased the number of activated microglia and astrocytes in the corpus callosum of MCAO rats. In addition, Western blot analysis indicated that CIG increased the expression of BDNF/p-TrkB, NRG1/ErbB4 proteins, which further elevated PI3K p110α/p-Akt/p-mTOR signaling in the corpus callosum of MCAO rats. CONCLUSION: We demonstrated that CIG protects against white matter lesions induced by cerebral ischemia partially by decreasing the number of activated microglia and astrocytes, increasing BDNF level, and activating NRG1/ErbB4 and its downstream PI3K/Akt/mTOR pathways in the white matter. CIG might be used as a potential neuroprotective agent for the treatment of ischemic stroke.

Probiotics Biofilm-Integrated Electrospun Nanofiber Membranes: A New Starter Culture for Fermented Milk Production.

Electrospun nanofiber membranes are widely investigated in the past few decades as candidates for tissue engineering, which can mimic natural extracellular matrix (ECM) and improve cell adhesion, proliferation, and expression on nanofiber membranes. However, the formation of bacterial biofilms on nanofiber membranes and application of the biofilm-integrated nanofiber membranes remain largely unknown. Here, electrospun cellulose acetate nanofiber membranes are first utilized as scaffold materials for Lactobacillus plantarum ( L. plantarum) biofilm formation. Nanofiber membranes proved to be an excellent scaffold for bacteria biofilm with high stability, where biofilms were interlocked with nanofibers forming a cohesive structure. In comparison with planktonic bacteria, L. plantarum biofilms on nanofiber membranes show excellent gastrointestinal resistance. Instead of decreasing, the number of viable cells increased after 3 h digestion in vitro. The L. plantarum biofilm-integrated nanofiber membranes were used as reusable starter cultures for fermented milk production showing excellent fermentative ability and higher survival of L. plantarum during shelf life. The viable cells in fermented milk remained at 11 log CFU/g throughout the reusable batches, which is far above the required value of 7 log CFU/g in commercial products. In addition, the produced fermented milk possesses shorter fermentation time and higher survival of probiotics during shelf life. The results suggest electrospun nanofiber membranes are ideal scaffold materials for bacteria biofilms immobilization in biotechnology and fermentation engineering, which broaden the potential use of electrospun nanofiber membranes in microbiology and strengthen the application of biofilms in fermentation engineering.

Atorvastatin improves coronary flow and endothelial function in patients with coronary slow flow.

The underlying mechanisms behind the effect of atorvastatin on patients with coronary slow flow (CSF) remain largely unknown. To investigate the possible underlying molecular mechanisms 108 patients were divided into atorvastatin group and control group. Coronary flow was quantified according to corrected TIMI frame count (CTFC). Serum high sensitivity C-reactive protein (hs-CRP), lipids, ET-1, interleukin (IL)-6, NO, circulating endothelial progenitor cell (cEPC) count, adhesion, migration and proliferation were measured in pretreatment and post-treatment. After respective treatment, the atorvastatin group had significantly decreased levels of TC, TG, LDL-C, hs-CRP, ET-1 and IL-6 and increased NO compared to the control group. The atorvastatin group had a more significant improvement of CTFC, effective rate, cEPC number, EPC adhesion, migration and proliferation compared to the control group. In conclusion, atorvastatin can be used in treatment of CSF by suppressing inflammation and improving endothelial function.

Substituting one Paris for another? In vitro cytotoxic and in vivo antitumor activities of Paris forrestii, a substitute of Paris polyphylla var. yunnanensis.

ETHNOPHARMACOLOGICAL RELEVANCE: Chong-lou (Paris polyphylla var. yunnanensis or P. polyphylla var. chinensis) is traditionally used as an anticancer medicine in China. It is also the material basis of some Chinese patent anticancer medicines, such as Gan-Fu-Le capsules, Bo-Er-Ning capsules, Lou-Lian capsules, Ruan-Jian oral liquid, and Qi-Zhen capsules. P. forrestii, a substitute for Chong-lou, is planted at a large scale in the Yunnan Province of China. AIM OF THE STUDY: To clarify the active chemical constituents of P. forrestii and evaluate the in vitro and in vivo anticancer activities of the total saponins from P. forrestii. MATERIALS AND METHODS: The total saponins of P. forrestii were extracted and separated to yield pure compounds by chromatographic techniques, and the structures of the isolates were elucidated by spectroscopic methods. The cytotoxicity of the crude extracts, total saponins, and chemical constituents were evaluated using an MTS assay. In vivo antitumor activities of the total saponins from P. forrestii were measured using H22 tumor-bearing mice by intraperitoneal (ip) administration. RESULTS: Eight compounds, including polyphyllin D (1), formosanin C (2), dioscin (3), diosgenin-3-O-α-l-rhamnopyranosyl-(1→2)-ß-d-glucopyranoside (4), paris saponin H (5), pennogenin-3-O-α-l-rhamnopyranosyl-(1→2)-[α-l-rhamnopyranosyl-(1→4)]-ß-d-glucopyranoside (6), pariposide A (7), and crustecdysone (8), were isolated from the total saponins of P. forrestii. The total saponins and compounds 1-6 showed significant inhibitory activity against the growth of the HL-60, SMMC-7721, A-549, MCF-7, and SW480 cell lines. The total saponins from P. forrestii had a tumor-inhibitory effect in H22 tumor-bearing mice upon ip (2.25 mg/kg dose) administration, with an inhibition rate of 42.6% compared with cisplatin (ip, 2 mg/kg dose, 53.9% inhibition rate). CONCLUSION: The results support that P. forrestii could be a substitute for P. polyphylla var. yunnanensis as an anticancer medicine.

A biotin-drug extraction and acid dissociation (BEAD) procedure to eliminate matrix and drug interference in a protein complex anti-drug antibody (ADA) isotype specific assay.

Monitoring anti-drug antibody (ADA) responses in patients receiving protein therapeutics treatment is an important safety assessment for regulatory agencies, drug manufacturers, clinicians and patients. Recombinant human IGF-1/IGFBP-3 (rhIGF-1/rhIGFBP-3) is a 1:1 formulation of naturally occurring protein complex. The individual IGF-1 and IGFBP-3 proteins have multiple binding partners in serum matrix with high binding affinity to each other, which presents challenges in ADA assay development. We have developed a biotin-drug extraction with acid dissociation (BEAD) procedure followed by an electrochemiluminescence (ECL) direct assay to overcome matrix and drug interference. The method utilizes two step acid dissociation and excess biotin-drug to extract total ADA, which are further captured by soluble biotin-drug and detected in an ECL semi-homogeneous direct assay format. The pre-treatment method effectively eliminates interference by serum matrix and free drug, and enhances assay sensitivity. The assays passed acceptance criteria for all validation parameters, and have been used for clinical sample Ab testing. This method principle exemplifies a new approach for anti-isotype ADA assays, and could be an effective strategy for neutralizing antibody (NAb), pharmacokinetic (PK) and biomarker analysis in need of overcoming interference factors.

[Medicinal values and their chemical bases of Paris].

Medicinal values and their chemical bases of Paris (Trilliaceae) are reviewed. Paris plants include 40 species and varieties. Among them, 18 ones are medicinal plants with similarity in traditional uses. Fourteen species have been studied phytochemically, which led to isolation of 207 compounds including 121 steroidal saponins. These saponins are major active constituents from Paris plants, which can explain the traditional uses of the plants to treat cancer, malignant boil, bleeding, gastritis, and so on. The similarity in medicinal uses and chemical constituents of Paris plants implies the possibility of resource substitution among these species. It is worth to further investigate Paris plants in chemical constituents, pharmacological activity, biological property, and toxicology.

Anticancer principles from medicinal piper ( hú jiao) plants.

The ethnomedical uses of Piper ( Hú Jiao) plants as anticancer agents, in vitro cytotoxic activity of both extracts and compounds from Piper plants, and in vivo antitumor activity and mechanism of action of selected compounds are reviewed in the present paper. The genus Piper (Piperaceae) contains approximately 2000 species, of which 10 species have been used in traditional medicines to treat cancer or cancer-like symptoms. Studies have shown that 35 extracts from 24 Piper species and 32 compounds from Piper plants possess cytotoxic activity. Amide alkaloids account for 53% of the major active principles. Among them, piplartine (piperlongumine) shows the most promise, being toxic to dozens of cancer cell lines and having excellent in vivo activity. It is worthwhile to conduct further anticancer studies both in vitro and in vivo on Piper plants and their active principles.

Diarylheptanoids and phenylphenalenones from Musa itinerans fruits.

Two diarylheptanoids, musaitinerins A and B, one heterodimeric phenylphenalenone musaitinerone and four known phenylphenalenones, identified as 4-hydroxy-2-methoxy-9-phenyl-1H-phenalen-1-one, musanolone E, hydroxyanigorufone and irenolone were isolated from the fruits of Musa itinerans Cheesm. Their structures were elucidated using spectroscopic analyses. The antimicrobial activity of these compounds was evaluated against Escherichia coli, Staphylococcus aureus and Candida albicans; the cytotoxic activity of these compounds was also evaluated against human erythromyeloblastoid leukemia (K562) and human alveolar carcinoma epithelial (A549) cell lines, respectively. Musaitinerone and musanolone E exhibited weak effects against the A549 cell line, as compared with adriamycin. However, these two compounds did not exhibit any growth inhibition against K562 cells, S. aureus, E. coli or C. albicans. The other compounds were inactive against all of the tested cell lines and microorganisms, even at concentrations as high as 50 µM.