Non-acid Components Research Articles

The Protease Inhibitor Amprenavir Protects against Pepsin-Induced Esophageal Epithelial Barrier Disruption and Cancer-Associated Changes.

Gastroesophageal reflux disease (GERD) significantly impacts patient quality of life and is a major risk factor for the development of Barrett's esophagus (BE) and esophageal adenocarcinoma (EAC). Proton pump inhibitors (PPIs) are the standard-of-care for GERD and are among the most prescribed drugs in the world, but do not protect against nonacid components of reflux such as pepsin, or prevent reflux-associated carcinogenesis. We recently identified an HIV protease inhibitor amprenavir that inhibits pepsin and demonstrated the antireflux therapeutic potential of its prodrug fosamprenavir in a mouse model of laryngopharyngeal reflux. In this study, we assessed the capacity of amprenavir to protect against esophageal epithelial barrier disruption in vitro and related molecular events, E-cadherin cleavage, and matrix metalloproteinase induction, which are associated with GERD severity and esophageal cancer. Herein, weakly acidified pepsin (though not acid alone) caused cell dissociation accompanied by regulated intramembrane proteolysis of E-cadherin. Soluble E-cadherin responsive matrix metalloproteinases (MMPs) were transcriptionally upregulated 24 h post-treatment. Amprenavir, at serum concentrations achievable given the manufacturer-recommended dose of fosamprenavir, protected against pepsin-induced cell dissociation, E-cadherin cleavage, and MMP induction. These results support a potential therapeutic role for amprenavir in GERD recalcitrant to PPI therapy and for preventing GERD-associated neoplastic changes.

Are the typical organic components in biomass pyrolyzed bio-oil available for leaching of alkali and alkaline earth metallic species (AAEMs) from biomass?

In this study, six dominant organic compounds in biomass pyrolyzed bio-oil, composed of acidic compound (acetic acid) and non-acidic compound (furfural, hydroxyacetone, ethylene glycol, phenol, and guaiacol) were used to remove AAEMs from different lignocellulosic biomass (rice husk, cotton stalk, and fir sawdust) by leaching pretreatment. Other four conventional solutions (deionized water, hydrochloric acid solution, aqueous phase bio-oil, and simulated bio-oil) were selected as control eluents. Results showed that the removal rate of AAEMs was highly affected by the types of immersion solutions, pH values, and the species of biomass. Majority of K in biomass could be removed by leaching of acetic acid. Ca was more difficult to be removed by leaching of water and acetic acid, but could be removed by leaching of HCl. In general, the removal rate of K was highest among the four species of AAMEs by using five non-acidic components, followed by Na, Mg, and Ca. The use of five non-acidic solutions was significantly benefit for the remove of K, Mg, and Na, but, was less useful in removal of Ca in rice husk and fir sawdust. However, the removal rates of Ca in cotton stalk by leaching of phenol, guaiacol, and hydroxyacetone were increased to nearly three times than water leaching. A synergistic effect for removing AAEMs occurred between acetic acid and non-acidic components in bio-oil, indicating that the non-acidic compounds also play an important role in leaching of AAEMs.

Experimental study on a biomass-based catalyst for catalytic upgrading and viscosity reduction of heavy oil

A biomass-based catalyst for catalytic upgrading and viscosity reduction process of heavy oil was prepared. The experimental results showed the viscosity of oil sample reduced from 508800 mPa.s to 22230 mPa.s (50 ℃) with 0.2wt% biomass-based catalyst at 320 ℃ for 30 min, and the viscosity reduction was up to 95.63%. The content of resin was significantly reduced by 10.1% after reaction under the optimal conditions. Furthermore, composition analysis, FT-IR analysis, thermogravimetric analysis and elemental analysis of the biomass-based catalyst were characterized in detail. In addition, FT-IR analysis, elemental analysis and SEM analysis were employed to evaluate the properties of oil sample. Further, influence mechanism of biomass-based catalyst on catalytic upgrading process of heavy oil was expounded through viscosity change, SARA analysis, 1H-NMR analysis, weight average molecular weight and average particle sizes analysis, and the above analysis results revealed that under the catalytic action of organic nickel salt, the presence of non-acidic components in the catalyst could improve the chemical structures of heavy compositions of oil sample. Finally, a possible schematic diagram of catalytic upgrading process was simply depicted in this work.

Airway reflux.

An ever-increasing number of adult and pediatric disorders have been shown to be influenced or caused by airway reflux. This has become a controversial and complicated aspect of medicine that requires a multidisciplinary approach. Evidence indicates that it is not only the acidic components of gastric refluxate that injure extraesophageal tissues but also the nonacidic components, such as pepsin and bile. There is a realization that proton pump inhibitors will not be effective when nonacidic components of refluxate are causing the problem. New in vitro and in vivo models for the study of airway reflux and new therapeutic and surgical approaches are discussed in this review article.

Bioassay guided identification of small chaperone proteins α-crystallin and Hsp27 inhibitors from Copaiba oil

Over-expression of small chaperone proteins in cancer tissue contributes to the resistance of chemotherapy. Targeting these small chaperones including α-crystallin and heat shock protein 27 (Hsp27) is a promising strategy for cancer treatment. Hardwickiic acid (HA), a clerodane diterpenoid from Copaiba oil has been reported to inhibit Hsp27. We expect to identify new small chaperone inhibitors from Copaiba oil that is abundant of diterpenoids. In the current study, cytotoxicity and anti-chaperone assay guided isolation of Copaiba oil led to two major fractions (non-acidic and acidic components) and seven sub-fractions from the acidic components. The non-acidic components and one sub-fraction showed significant cytotoxicity in prostate cancer cells. Four sub-fractions exhibited potent anti-chaperone activity. Three chemical components were identified from these sub-fractions including copalic acid, hardwickiic acid and 3-acetoxycopalic acid. All three compounds inhibited the chaperone activities of α-crystallin and Hsp27. In addition, these compounds enhanced the anti-proliferative activity of the chemotherapeutic agent carboplatin in LNCaP cells. 3-Acetoxycopalic acid slightly decreased the level of Hsp27 client protein signal transducer and activator of transcription 3 (Stat3) in PC3 cells. Overall, several diterpenoids were identified to be small chaperone inhibitors and could be used as lead compounds for the development of more potent derivatives.

Prevalence of gastroesophageal reflux in cystic fibrosis and implications for lung disease.

Gastroesophageal reflux (GER) is common in patients with cystic fibrosis (CF) and is often regarded as playing a role in the pathogenesis of CF lung disease. Individuals with CF have many predisposing factors to the development of GER, with a reported prevalence ranging from 35 to 81%. Several studies have suggested that patients with CF who have coexisting GER have more severe lung disease with lower pulmonary function and increased numbers of respiratory exacerbations. Furthermore, GER may alter the respiratory microbiology in CF. Both the acid and nonacid components of GER may have an effect on lung disease. More than 50% of U.S. patients with CF were being treated with proton pump inhibitors in 2012; however, data regarding safety and efficacy of these agents in CF are lacking. Pharmacologic and surgical treatment of GER may improve respiratory morbidity, although prospective controlled studies have not been performed. Given the lack of evidence-based guidelines for evaluation, diagnosis, and treatment of GER in CF, initiation of treatment for symptomatic GER should be based on standard guidelines for the general population. Because there is no clear evidence that GER leads to worse respiratory outcomes in CF or that treatment of GER improves pulmonary outcomes, invasive testing for GER in patients without reflux symptoms is not warranted. Further studies to determine the role of GER in CF lung disease and the risks and benefits of surgical and pharmacologic therapy for GER are warranted.

Su1252 Gastric Juice Composition and Acid Suppression in Pediatric Gastroesophageal Reflux Disease

INTRODUCTION: Gastric acid suppression is justified to prevent severe gastro esophageal reflux (GER) disease related complications. However, it does not reduce the total amount and proximal extent of GER in the esophagus and non-acid components are able to induce (extra-esophageal) GER symptoms as well. No data on the composition of gastric juice (GJ) in children using gastric acid suppression exists. We therefore aimed to assess whether the composition of gastric juice in children using proton pump inhibitors (PPIs) differs, compared to that of their controls. METHODS: Infants and children (0-18 years) on proton pump inhibitors (PPIs) for at least six weeks and a control group not using anti reflux medication, were included. GJ was obtained through an existing nasogastric or a percutaneous endoscopic gastrostomy tube/ Mic-key gastrostomy. In the collected GJ (5 ml), pH, pepsin activity, bile salts and endotoxin (LPS) levels were determined. Pepsin was measured using a fluorometric assay using 4-Methyl-Coumaryl-7-Amide (MCA) substrate with/without pepstatin. Concentrations of deconjugated and taurine/glycine-conjugated bile salts were assessed by reversephase HPLC. Levels of LPS were determined using the spectrophotometric Limulus Amebocyte Lysate assay. RESULTS: GJ was analyzed from 16 children with (median: 3.8 yrs, range: 17.6 years) and 16 children (4.0 yrs, range: 16.0) without PPI therapy. Median duration of PPI treatment was 24 weeks (range: 514 weeks). Gastric pH was 5.0 (range: 5.0) and 1.0 (range: 4.5) in the PPI and control group respectively (p <0.001). Pepsin, unconjugated bile salts, and endotoxin were not significantly different in the two groups. Total taurine conjugated bile salts, and specifically taurocholate, was significantly higher in the PPI group (p=0.01 and p=0.005). pH and concentration of deconjugated bile and conjugated bile salts were significantly associated (p=0.006 and p=0.02). Endotoxin and bile salts were not significantly associated. CONCLUSION: Taurine conjugated bile acids are significantly higher in children chronically using PPIs compared to controls. Moreover acidity of gastric pH correlated negatively with deconjugated and conjugated bile salts. These findings imply that GJ under chronic proton pump inhibition contains nonacid components potentially harmful to esophageal mucosa and bronchial tissue.

Su1253 Abnormal Dynamics of Endogenous Acylated Ghrelin Mediates Decreases in Gastric Phase III-Like Contraction, Leading to Delayed Gastric Emptying in Urocortin1-Induced Stress Rats

INTRODUCTION: Gastric acid suppression is justified to prevent severe gastro esophageal reflux (GER) disease related complications. However, it does not reduce the total amount and proximal extent of GER in the esophagus and non-acid components are able to induce (extra-esophageal) GER symptoms as well. No data on the composition of gastric juice (GJ) in children using gastric acid suppression exists. We therefore aimed to assess whether the composition of gastric juice in children using proton pump inhibitors (PPIs) differs, compared to that of their controls. METHODS: Infants and children (0-18 years) on proton pump inhibitors (PPIs) for at least six weeks and a control group not using anti reflux medication, were included. GJ was obtained through an existing nasogastric or a percutaneous endoscopic gastrostomy tube/ Mic-key gastrostomy. In the collected GJ (5 ml), pH, pepsin activity, bile salts and endotoxin (LPS) levels were determined. Pepsin was measured using a fluorometric assay using 4-Methyl-Coumaryl-7-Amide (MCA) substrate with/without pepstatin. Concentrations of deconjugated and taurine/glycine-conjugated bile salts were assessed by reversephase HPLC. Levels of LPS were determined using the spectrophotometric Limulus Amebocyte Lysate assay. RESULTS: GJ was analyzed from 16 children with (median: 3.8 yrs, range: 17.6 years) and 16 children (4.0 yrs, range: 16.0) without PPI therapy. Median duration of PPI treatment was 24 weeks (range: 514 weeks). Gastric pH was 5.0 (range: 5.0) and 1.0 (range: 4.5) in the PPI and control group respectively (p <0.001). Pepsin, unconjugated bile salts, and endotoxin were not significantly different in the two groups. Total taurine conjugated bile salts, and specifically taurocholate, was significantly higher in the PPI group (p=0.01 and p=0.005). pH and concentration of deconjugated bile and conjugated bile salts were significantly associated (p=0.006 and p=0.02). Endotoxin and bile salts were not significantly associated. CONCLUSION: Taurine conjugated bile acids are significantly higher in children chronically using PPIs compared to controls. Moreover acidity of gastric pH correlated negatively with deconjugated and conjugated bile salts. These findings imply that GJ under chronic proton pump inhibition contains nonacid components potentially harmful to esophageal mucosa and bronchial tissue.

Abstract B168: Dietary polyphenols inhibit pepsin and prevent airway epithelial cell hyperproliferation associated with laryngeal cancer.

Abstract A number of otolaryngologic inflammatory and neoplastic diseases are associated with gastric reflux extending proximal to the esophagus. Extraesophageal reflux, also known as laryngopharyngeal reflux (LPR), increases the risk of developing laryngeal cancer by approximately two-fold and analyses of neoplastic development in patients with prior gastrectomy suggest that nonacid components of refluxate promote laryngeal carcinogenesis as well. Clinical and basic science research implicates gastric pepsin as a key deleterious agent of LPR. Our previous research demonstrates that pepsin at pH7 induces hyperproliferation and inflammatory and cell cycle regulatory gene expression changes in airway epithelial cells in vitro. Acid suppression therapy through proton-pump inhibitors (PPI's), the most commonly prescribed therapeutic for reflux, has demonstrated minimal efficacy in resolution of LPR-attributed disease. Phytochemicals derived from algae, pine, and most recently Noni fruit, however, have been shown to not only attenuate reflux-attributed lesions and inflammation in rat models of disease but also directly inhibit peptic activity. This study examines the potential of plant polyphenols curcumin, ecabet sodium (ES) and an anthocyanin-enriched fraction of freeze-dried black raspberry (BRB) to inhibit peptic activity and pepsin-mediated cell damage and proliferative changes in cultured human laryngeal and pharyngeal cells. Cultured human laryngeal epithelial cells from punch biopsy, pharyngeal carcinoma (FaDu) and gastric adenocarcinoma (AGS) cells were pre-treated with/without curcumin (10uM), ES (125ug/ml) and BRB (100ug/ml) for 30 minutes, rinsed and subsequently treated with/without nonacid pepsin (0.1mg/ml, pH7.0) for 1 hour. Morphological changes were examined by transmission electron microscopy and proliferative changes by cell count assay and realtime PCR array (curcumin only). Direct inhibition of peptic activity was determined by in vitro kinetic assay. Curcumin, ES and BRB significantly inhibited peptic activity (p&amp;lt;0.05) in a dose-dependent manner at uM or ug/ml concentrations. Curcumin, ES and BRB abrogated mitochondrial damage and significantly reduced hyperproliferation elicited by pepsin in human laryngeal and pharyngeal cells. Curcumin attenuated 28 of the 31 changes in cell cycle regulatory gene expression induced by pepsin in pharyngeal cells. Gastric cells, which would not be expected to exhibit sensitivity to pepsin-mediated damage, displayed neither mitochondrial damage nor hyperproliferation following treatment with pepsin. These data demonstrate that plant polyphenols curcumin, ES and BRB are protective against pepsin-mediated airway epithelial cell damage and hyperplasia and implicate dietary modification/supplementation with plant polyphenols as a novel pepsin-protective therapeutic modality for diseases associated with proximal reflux including laryngeal cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr B168.

Preparation and Evaluation of Epoxy Binders Based on Rosin as Organic Coating for Steel

Abstract: In this work, modification of rosin as epoxy organic coating was reported. Epoxy resins were prepared from modified rosin, rosin Diels-Alder adducts, rosin formaldehyde and their ketone. These adducts were reacted with epichlorohydrine in the presence of NaOH as a catalyst to produce epoxy resins. The preparation of the curing agents for the epoxy resins based on rosin poly (amideimide) was investigated. The recent advances in manipulating patterned rosin arrays and patents are reviewed with a focus on the progress of production of epoxy resins based on biopolymer and natural products and their epoxy organic coating applications. Keywords: Rosin, corrosion protection, organic coatings, epoxy resins, coating characteristics. 1. INTRODUCTION Epoxy resins have been widely used in many industrial applications because of their good heat and chemical resistance, superior mechanical and electrical properties and excellent process capabilities [1]. Intensive research efforts have been devoted to develop new epoxy resins through chemical and physical modifications of the classical resins in the last few years [2, 3]. It was also noted that the epoxy resin based paints generally offer very good corrosion protective properties and are widely used for different applications. Rosin is a complex mixture of mainly twenty carbon atom fused rings, mono-carboxylic acids and a small amount of nonacidic components, where the resin acid molecule has double bonds and the carboxylic acid group, which may be possible for a derivative to be used that maintains the carboxylic acid group. Several patents used rosin in epoxy resin formulation as linking agents [4-8]. Modified epoxy resins comprising the reaction product of rosin and a linking molecule reacted with an epoxy resin are disclosed. Aqueous dispersions and coatings comprising these resins are also reported [4, 5]. On the other hand, rosin was modified and used as a solid curing agent in a composition for coating a substrate with a powder coating comprising a solid polyepoxide resin [6]. Rosin as crosslinking agent is essentially free of active hydrogen functional groups. Methods for preparing rosin as crosslinking agents for aminoplast formulation were investigated

The role of an alginate suspension on pepsin and bile acids - key aggressors in the gastric refluxate. Does this have implications for the treatment of gastro-oesophageal reflux disease?

During a reflux event the oesophagus is exposed to a heterogeneous mixture of gastric juice components. The role of non-acid components of the refluxate in causing damage to the oesophagus is now well established but no therapeutic option exists to address this. The role of Gaviscon Advance (GA), a raft-forming alginate suspension, in protecting the oesophagus from damage by pepsin and bile acids (aggressors) was investigated using a series of in-vitro models. GA was able to dose-dependently inhibit pepsin activity over and above the neutralisation effect of the formulation. This was evident against both protein and collagen substrates using two distinct colorimetric assays. GA was able to retard the diffusion of pepsin and multiple bile acids using a Franz cell model. Using the raft-forming mode of action GA was able to remove both pepsin and multiple bile acids from a simulated reflux event. There was capacity in the GA raft to accommodate aggressors from multiple reflux events. GA can specifically remove both pepsin and bile acids from the refluxate, limit their diffusion and affect enzymatic activity of pepsin. There is a role for GA to reduce the damaging potential of the refluxate and thus protect the oesophagus.

The role of an alginate suspension on pepsin and bile acids – key aggressors in the gastric refluxate. Does this have implications for the treatment of gastro-oesophageal reflux disease?

Abstract Objectives During a reflux event the oesophagus is exposed to a heterogeneous mixture of gastric juice components. The role of non-acid components of the refluxate in causing damage to the oesophagus is now well established but no therapeutic option exists to address this. Methods The role of Gaviscon Advance (GA), a raft-forming alginate suspension, in protecting the oesophagus from damage by pepsin and bile acids (aggressors) was investigated using a series of in-vitro models. Key findings GA was able to dose-dependently inhibit pepsin activity over and above the neutralisation effect of the formulation. This was evident against both protein and collagen substrates using two distinct colorimetric assays. GA was able to retard the diffusion of pepsin and multiple bile acids using a Franz cell model. Using the raft-forming mode of action GA was able to remove both pepsin and multiple bile acids from a simulated reflux event. There was capacity in the GA raft to accommodate aggressors from multiple reflux events. Conclusions GA can specifically remove both pepsin and bile acids from the refluxate, limit their diffusion and affect enzymatic activity of pepsin. There is a role for GA to reduce the damaging potential of the refluxate and thus protect the oesophagus.

Review article: the laryngological manifestations of reflux disease; why the scepticism?

Despite increasing clinical and experimental evidence of its existence, otolaryngological manifestations of reflux disease remain controversial, concerning diagnosis as well as treatment. Proper understanding of laryngopharyngeal reflux disease (LPRD). Review of literature. Scepticism concerning LPRD is based upon differences between gastro-oesophageal reflux disease and LPRD; lack of specificity and sensitivity of diagnostic tests to confirm LPRD; non-specificity of laryngological symptoms, which are difficult to distinguish from other causes of upper respiratory tract inflammation; non-specificity of laryngological signs in laryngoscopy, with high intra- and inter-observer variability in evaluation; diagnosis of LPRD is essentially only based on a combination of diagnostic signs and symptoms, which cannot be attributed to other pathology; slow, or sometimes lack of, response of LPRD symptoms to proton pump inhibitor (PPI) medication and lack of evidence concerning efficacy of PPIs in placebo-controlled trials. LPRD remains a diagnosis by exclusion and resolution of symptoms following 4-month trial of 40 mg PPI twice daily is, for all practical purposes, considered proof of the initial diagnosis. However, non-response does not exclude LPRD as PPIs have no influence on noxious non-acid components of the refluxate.

Laryngopharyngeal Reflux: Beauty is in The Eye of The Beholder

Laryngopharyngeal Reflux: Beauty is in The Eye of The Beholder

Development of an odour‐baited target for female New World screwworm, Cochliomyia hominivorax: studies with host baits and synthetic wound fluids

Field studies were conducted in Mexico to investigate the possibility of replacing sentinel animals as baits for female New World screwworm Cochliomyia hominivorax (Coquerel) (Diptera: Calliphoridae) with a synthetic odour-bait composed of compounds identified from wounds infested with the larvae of C. hominivorax. Studies using sheep held in stanchions surrounded by an incomplete ring of electrified nets showed that both male and female C. hominivorax were attracted to an unwounded animal (ratio of male : female flies caught 1.11 : 1.00), although they were not observed to land. By contrast, catches associated with unwounded sheep with larval wound fluid applied to shaven shoulders suggested that female C. hominivorax were attracted to (male : female ratio of electrified nets catch 0.14 : 1.00) and landed on the wound fluid but male flies did not respond (male : female ratio alighting on fluid 0.05 : 1.00). In related studies the attractiveness of synthetic blends of 25 electrophysiologically active compounds identified in fluids associated with wounds infested with the larvae of C. hominivorax were tested. The acidic components of the wound fluids were found to attract and elicit a landing response from both male and female flies, whereas the non-acidic components alone caught only low numbers of flies. However, the numbers of male and female flies that were attracted to and landed on a synthetic bait could be increased significantly by increasing the proportion of non-acid to acid components in the lure. In some replicates the most effective blend caught a number of C. hominivorax comparable to that caught by a standard synthetic attractant, Swormlure-4, although, unlike with Swormlure-4, the catch was predominately composed of female flies. It is uncertain whether Swormlure-4 contains compounds that elicit a landing response from male C. hominivorax or whether the synthetic and natural wound odours contain compounds that inhibit a landing response from male C. hominivorax. Further work is required to simplify the blend of compounds needed to attract female flies and to better understand the role of the compounds that elicit behavioural responses from both male and female C. hominivorax before a synthetic substitute can be fully developed to replace sentinel animals.

Review article: the role of bile and pepsin in the pathophysiology and treatment of gastro‐oesophageal reflux disease

Gastro-oesophageal reflux disease is a multifaceted and multifactorial disorder which results from the reflux of gastric contents into the oesophagus. Animal studies suggest that synergism between acid and pepsin and conjugated bile acids have the greatest damaging potential for oesophageal mucosa, although unconjugated bile acids may be caustic at more neutral pH. Human studies are compatible with a synergistic action between acid and duodenogastric reflux in inducing lesions. During prolonged monitoring studies, typical gastro-oesophageal reflux symptoms are more related to acid reflux events than to non-acid reflux events. However, symptoms that persist during acid suppressive therapy are often related to non-acid reflux events. The therapeutic options for the non-acid component of the refluxate, including acid suppression, prokinetics, baclofen, surgery and mucosal protective agents like alginates, are discussed.

Review article: role of pepsin and bile in gastro‐oesophageal reflux disease

Gastro-oesophageal reflux disease is defined as the presence of symptoms or lesions that can be attributed to the reflux of gastric contents into the oesophagus. Aspiration and prolonged monitoring studies in humans have shown that reflux of gastric contents is comprised of both acid and non-acid components, in healthy as well as diseased people. Methods to monitor the non-acid component of the refluxate are described in detail. Experimental models suggest that synergism between acid and pepsin and conjugated bile acids have the greatest damaging potential for oesophageal mucosa, although unconjugated bile acids may be caustic at a more neutral pH. Human studies are compatible with a synergistic action between acid and duodenogastric reflux in inducing lesions. During prolonged monitoring studies, typical gastro-oesophageal reflux disease symptoms are more related to acid reflux events than to non-acid reflux events. However, symptoms that persist during acid-suppressive therapy are often related to non-acid reflux events. The therapeutic options for the non-acid component of the refluxate, including acid suppression, prokinetics, baclofen and surgery, are discussed.

Cytotoxic effects of current dental adhesive systems on immortalized odontoblast cell line MDPC-23

Objectives: Evaluate the cytotoxic effect of the three dental adhesive systems. Methods: The immortalized mouse odontoblast cell line (MDPC-23) was plated (30,000 cell/cm 2) in 24 well dishes, allowed to grow for 72 h, and counted under inverted light microscopy. Uncured fresh adhesives were added to culture medium to simulate effects of unset adhesive. Three adhesives systems were applied for 120 min to cells in six wells for each group: Group 1) Single Bond (3M), Group 2) Prime & Bond 2.1 (Dentsply), and Group 3) Syntac Sprint (Vivadent). In the control group, PBS was added to fresh medium. The cell number was counted again and the cell morphology was assessed under SEM. In addition, the adhesive systems were applied to circles of filter paper, light-cured for 20 s, and placed in the bottom of 24 wells (six wells for each experimental materials and control group). MDPC-23 cells were plated (30,000 cell/cm 2) in the wells and allowed to incubate for 72 h. The zone of inhibition around the filter papers was measured under inverted light microscopy; cell morphology was evaluated under SEM; and the MTT assay was performed for mitochondrial respiration. Results: The fresh adhesives exhibited more toxic (cytopathic effects) to MDPC-23 cells than polymerized adhesives on filter papers, and as compared to the control group. The cytopathic effect of the adhesive systems occurred in the inhibition zone around the filter papers, which was confirmed by the MTT assay and statistical analysis (ANOVA) combined with Fisher's PLSD test. In the control group, MDPC-23 cells were dense on the plastic substrate and were in contact with the filter paper. In the experimental groups, when acid in the adhesive systems was removed by changing the culture medium, or when the adhesives were light-cured, some cells grew in the wells in spite of the persistent cytotoxic effect. Significance: All dentin adhesive systems were cytotoxic odontoblast-like cells. Both acidity and non-acidic components of these systems were responsible for the high cytopathic effect of those dental materials.

Analysis of methyl tert.-butyl ether and its degradation products by direct aqueous injection onto gas chromatography with mass spectrometry or flame ionization detection systems

A method was developed to analyze methyl tert.-butyl ether (MTBE) and its degradation products by gas chromatography with mass spectrometry (GC–MS) or flame ionization detection (FID) with direct aqueous injection. The column had dimensions of 30 m×0.25 mm with film thickness 0.25 μm and a stationary phase of FFAP (nitroterephthalic acid-modified polyethylene glycol). The optimized GC conditions for non-acid components were as follows: carrier gas flow-rate,1 mL/min; oven temperature, 35°C for 5.5 min, ramped to 90°C at 25°C/min, then ramped to 200°C at 40°C/min and held at 200°C for 8 min. The conditions for the acid components were: carrier gas flow-rate, 1 mL/min; oven temperature, 110°C for 2 min, ramped to 150°C at 10°C/min, then ramped to 200°C at 40°C/min. The injection port contained a silanized-glass reverse-cup liner filled with Carbofrit. The minimum concentrations for the linear range for the selective ion monitoring mode were 30 to 100 μg/L, depending on the analytes. The minimum detection limit was 1 mg/L for MTBE and tert.-butanol when using FID. More components could be analyzed with the FFAP-type column than with the cyanopropylphenyl–dimethyl polysiloxane-type column.

Identification of electrophysiologically-active compounds for the malaria mosquito, Anopheles gambiae, in human sweat extracts.

Human sweat samples were chemically fractionated into acid and non-acid components. The most abundant volatile compounds present in the fractions were identified by linked gas chromatography mass spectrometry. The acid fractions were found to be composed of a range of twenty aliphatic and three aromatic carboxylic acids ranging, on average, from 0.02 to 20 micrograms per ml of sweat sampled. Non-acid fractions were found to contain: 6-methyl-5-hepten-2-one, 1-octen-3-ol, decanal, benzyl alcohol, dimethylsulphone, phenylethanol, phenol and 4-methylphenol, collectively amounting to 0.1 and 3 micrograms per ml of sweat. The major component of sweat was found to be L-lactic acid which constituted from 1 to 5 mg/ml. Using the intact antennae of the anthropophilic malaria vector mosquito Anopheles gambiae Giles, the peripheral olfactory activities of compounds identified in the sweat fractions were investigated by electroantennography (EAG). Short-chain saturated carboxylic acids, methanoic, ethanoic, propanoic, butanoic, pentanoic and hexanoic acids were found to elicit significantly larger EAG responses than longer chain saturated carboxylic acids from female An.gambiae. For a given dose the largest amplitude EAG response was elicited by methanoic acid. Pentanoic acid elicited larger EAG responses than either butanoic or hexanoic acids. Two non-acidic compounds, 1-octen-3-ol and 4-methylphenol, were found to elicit significant dose-dependent EAG responses from female An.gambiae. 1-Octen-3-ol elicited larger EAG responses than 4-methylphenol for a given dose, but both compounds elicited smaller EAG responses than the same dose of C1-C6 straight-chain aliphatic carboxylic acids. The possible behavioural significance of the EAG-active compounds identified in human sweat samples is discussed.