Acrolein synthesis from glycerol in hot-compressed water

Glycerol conversion was conducted in hot-compressed water (HCW: 573-673 K, 25-34.5 MPa) using a batch and a flow apparatus and the influences of temperature, H(2)SO(4), glycerol concentration, and pressure, were examined. The yield of acrolein was enhanced by higher glycerol and H(2)SO(4) concentration, and higher pressure. Approximately 80% selectivity of acrolein was obtained at 90% of glycerol conversion with an acid catalyst in supercritical condition (673 K and 34.5 MPa). The rate constant of acrolein decomposition was always higher than that of acrolein formation in the absence of acid catalyst but the rate constant of acrolein formation could be overcome that of acrolein decomposition by addition acid in supercritical condition.     

Conversion of allyl alcohol into acrolein by rat liver

1. Acrolein was detected in rat liver suspensions incubated in the presence of allyl alcohol and allyl formate. Acrolein was determined by condensation of the distilled aldehyde with semicarbazide, followed by spectrophotometric measurement of the semicarbazone at 257nm and identification by paper chromatography. 2. The transformation of allyl alcohol into acrolein occurred in the presence of NAD(+). Inhibitors of rat liver alcohol dehydrogenase inhibited the reaction. 3. It is suggested that the hepatotoxic actions of allyl alcohol and of allyl formate on rat liver are related to their conversion into acrolein.     

The antihypertensive hydralazine is an efficient scavenger of acrolein

Recent work indicates the highly toxic alpha,beta-unsaturated aldehyde acrolein is formed during the peroxidation of polyunsaturated lipids, raising the possibility that it functions as a 'toxicological second messenger' during oxidative cell injury. Acrolein reacts rapidly with proteins, forming adducts that retain carbonyl groups. Damage by this route may thus contribute to the burden of carbonylated proteins in tissues. This work evaluated several amine compounds with known aldehyde-scavenging properties for their ability to attenuate protein carbonylation by acrolein. The compounds tested were: (i) the glycoxidation inhibitors, aminoguanidine and carnosine; (ii) the antihypertensive, hydralazine; and (iii) the classic carbonyl reagent, methoxyamine. Each compound attenuated carbonylation of a model protein, bovine serum albumin, during reactions with acrolein at neutral pH and 37 degrees C. However, the most efficient agent was hydralazine, which strongly suppressed carbonylation under these conditions. Study of the rate of reaction between acrolein and the various amines in a protein-free buffered system buttressed these findings, since hydralazine reacted with acrolein at rates 2-3 times faster than its reaction with the other scavengers. Hydralazine also protected isolated mouse hepatocytes against cell killing by allyl alcohol, which undergoes in situ alcohol dehydrogenase-catalysed conversion to acrolein.     

Production of biodiesel and lactic acid from rapeseed oil using sodium silicate as catalyst

Biodiesel and lactic acid from rapeseed oil was produced using sodium silicate as catalyst. The transesterification in the presence of the catalyst proceeded with a maximum yield of 99.6% under optimized conditions [3% (w/w) sodium silicate, methanol/oil molar ratio 9/1, reaction time 60 min, reaction temperature 60 °C, and stirring rate 250 rpm]. After six consecutive transesterification reactions, the catalyst was collected and used for catalysis of the conversion of glycerol to lactic acid. A maximum yield of 80.5% was achieved when the reaction was carried out at a temperature of 300 °C for 90 min. Thus, sodium silicate is an effective catalyst for transesterification and lactic acid production from the biodiesel by-product, glycerol.     

Acrolein toxicity involves oxidative stress caused by glutathione depletion in the yeast <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

Exposure of yeast cells to allyl alcohol results in intracellular production of acrolein. The toxicity of so formed acrolein involves oxidative stress, as (1) strains deficient in antioxidant defense are hypersensitive to allyl alcohol, (2) exposure to allyl alcohol increases the level of thiobarbituric-acid-reactive substances and decreases glutathione level in the cells, (3) hypoxic and anoxic atmosphere and antioxidants protect against allyl alcohol toxicity, and (4) allyl alcohol causes activation of Yap1p. No increased formation of reactive oxygen species was detected in cells exposed to allyl alcohol, so oxidative stress is due to depletion of cellular thiols and thus alteration in the redox state of yeast cells.     

Etherification of biodiesel-derived glycerol with ethanol for fuel formulation over sulfonic modified catalysts

The present study is focused on the etherification of biodiesel-derived glycerol with anhydrous ethanol over arenesulfonic acid-functionalized mesostructured silicas to produce ethyl ethers of glycerol that can be used as gasoline or diesel fuel biocomponents. Within the studied range, the best conditions to maximize glycerol conversion and yield towards ethyl-glycerols are: T = 200 °C, ethanol/glycerol molar ratio = 15/1, and catalyst loading = 19 wt%. Under these reaction conditions, 74% glycerol conversion and 42% yield to ethyl ethers have been achieved after 4 h of reaction but with a significant presence of glycerol by-products. In contrast, lower reaction temperatures (T = 160 °C) and moderate catalyst loading (14 wt%) in presence of a high ethanol concentration (ethanol/glycerol molar ratio = 15/1) are necessary to avoid the formation of glycerol by-products and maximize ethyl-glycerols selectivity. Interestingly, a close catalytic performance to that achieved using high purity glycerol has been obtained with low-grade water-containing glycerol.     

The molecular mechanisms of diallyl disulfide and diallyl sulfide induced hepatocyte cytotoxicity

Diallyl disulfide (DADS) and diallyl sulfide (DAS) are the major metabolites found in garlic oil and have been reported to lower cholesterol and prevent cancer. The molecular cytotoxic mechanisms of DADS and DAS have not been determined.The cytotoxic effectiveness of hydrogen versus allyl sulfides towards hepatocytes was found to be as follows: NaHS > DADS > DAS. Hepatocyte mitochondrial membrane potential was decreased and reactive oxygen species (ROS) and TBARS formation was increased by all three allyl sulfides. (1) DADS induced cytotoxicity was prevented by the H₂S scavenger hydroxocobalamin, which also prevented cytochrome oxidase dependent mitochondrial respiration suggesting that H₂S inhibition of cytochrome oxidase contributed to DADS hepatocyte cytotoxicity. (2) DAS cytotoxicity on the other hand was prevented by hydralazine, an acrolein trap. Hydralazine also prevented DAS induced GSH depletion, decreased mitochondrial membrane potential and increased ROS and TBARS formation. Chloral hydrate, the aldehyde dehydrogenase 2 inhibitor, however had the opposite effects, which could suggest that acrolein contributed to DAS hepatocyte cytotoxicity.     

Lipase-catalyzed transesterification of soybean oil for biodiesel production in <Emphasis Type="Italic">tert</Emphasis>-amyl alcohol

Lipase-catalyzed transesterification of soybean oil and methanol for biodiesel production in tert-amyl alcohol was investigated. The effects of different organic medium, molar ratio of substrate, reaction temperature, agitation speed, lipase dosage and water content on the total conversion were systematically analyzed. Under the optimal conditions identified (6 mL tert-amyl alcohol, three molar ratio of methanol to oil, 2% Novozym 435 lipase based on the soybean oil weight, temperature 40°C, 2% water content based on soybean oil weight, 150 rpm and 15 h), the highest biodiesel conversion yield of 97% was obtained. With tert-amyl alcohol as the reaction medium, the negative effects caused by excessive molar ratio of methanol to oil and the by-product glycerol could be reduced. Furthermore, there was no evident loss in the lipase activity even after being repeatedly used for more than 150 runs.     

Optimization of direct conversion of wet algae to biodiesel under supercritical methanol conditions

This study demonstrated a one-step process for direct liquefaction and conversion of wet algal biomass containing about 90% of water to biodiesel under supercritical methanol conditions. This one-step process enables simultaneous extraction and transesterification of wet algal biomass. The process conditions are milder than those required for pyrolysis and prevent the formation of by-products. In the proposed process, fatty acid methyl esters (FAMEs) can be produced from polar phospholipids, free fatty acids, and triglycerides. A response surface methodology (RSM) was used to analyze the influence of the three process variables, namely, the wet algae to methanol (wt./vol.) ratio, the reaction temperature, and the reaction time, on the FAMEs conversion. Algal biodiesel samples were analyzed by ATR-FTIR and GC-MS. Based on the experimental analysis and RSM study, optimal conditions for this process are reported as: wet algae to methanol (wt./vol.) ratio of around 1:9, reaction temperature and time of about 255 °C, and 25 min respectively. This single-step process can potentially be an energy efficient and economical route for algal biodiesel production.     

Thermal glycosylation and degradation reactions occurring at the reducing ends of cellulose during low-temperature pyrolysis

Thermal glycosylation and degradation reactions of cellulose (Avicel PH-101) were studied in the presence or absence of alcohols (glycerol, mannitol, 1,2,6-hexanetriol, 3-phenoxy-1,2-propanediol, and 1-tetradecanol) under N₂ at 60–280 °C. In the presence of glycerol (heating time, 10 min), the reducing ends were converted into glycosides when the temperature of the glycerol was >140 °C without the addition of any catalysts. A temperature of 140 °C is close to that required for the initiation of thermal polymerization (glycosylation). Although the conversion was only around 20% in the range of 140–180 °C, the reactivity increased above 200–240 °C where the thermal expansion of cellulose crystals is reported to become significant. Finally, all reducing ends were converted into glycosides at 260 °C. Such heterogeneous reactivity likely arose from the lower reactivities of the reducing ends in the crystalline region due to their lower accessibility to glycerol, although the reactivity in the non-crystalline region was similar to that of glucose. Alcohols that have a lower OH/C ratio did not react with the reducing ends, suggesting that the hydrophilicity of the alcohol was critical for the glycosylation reaction to proceed. The glycosylated cellulose samples were found to be significantly stabilized against pyrolytic coloration. The results of neat cellulose pyrolysis indicated that two competitive reactions, thermal glycosylation and degradation, formed a dark-colored substance at the reducing ends while the internal glucose units in the cellulose were comparatively stable.     

Allyl alcohol toxicity in isolated renal epithelial cells: protective effects of low molecular weight thiols

The toxicity of allyl alcohol was studied in freshly isolated renal epithelial cells prepared from male and female rats. Cells from female rats demonstrated a greater susceptibility to allyl alcohol toxicity as assessed by glutathione depletion and loss of cell viability. The sensitivity of female rat renal cells appears to relate to the higher activity of alcohol dehydrogenase found in the female rat kidney, which metabolizes allyl alcohol to the highly reactive aldehyde, acrolein. Pyrazole, an inhibitor of alcohol dehydrogenase, abolished the cytotoxic effects of allyl alcohol whereas inhibition of aldehyde dehydrogenase by disulfiram treatment was found to increase the sensitivity of renal cells to the effects of allyl alcohol. The toxicity of allyl alcohol was decreased by a number of treatments which resulted in increased levels of glutathione or other low molecular weight thiols. These results indicate that acrolein is the toxic metabolite responsible for the renal cell injury following exposure to allyl alcohol, and unless immediately inactivated acrolein interacts with critical nucleophilic sites of the cell and initiates cell injury. These studies demonstrate that freshly isolated kidney cells represent a convenient model system for studies of thiol-mediated protective mechanisms against toxic renal cell injury.     

Dual response surface-optimized process for feruloylated diacylglycerols by selective lipase-catalyzed transesterification in solvent free system

Feruloylated diacylglycerol (FDAG) was synthesized using a selective lipase-catalyzed the transesterification between ethyl ferulate and triolein. To optimize the reaction conversion and purity of FDAG, dual response surface was applied to determine the effects of five-level-five-factors and their reciprocal interactions on product synthesis. A total of 32 individual experiments were performed to study reaction temperature, reaction time, substrate molar ratio, enzyme loading, and water activity. The highest reaction conversion and selectivity towards FDAG were 73.9% and 92.3%, respectively, at 55 °C, reaction time 5.3 day, enzyme loading 30.4 mg/ml, water activity 0.08, and a substrate molar ratio of 3.7. Moreover, predicted values showed good validation with the experimental values when experiments corresponding to selected points on the contour plots were carried out.     

The hepatotoxic action of allyl formate

The hepatotoxic action of allyl formate on rat liver has been investigated. Biochemical changes can be detected in the liver cell many hours before the histological changes and it would appear that the toxin has a direct action on the liver parenchymal cell. The results suggest that allyl formate is not the toxic agent but that it is converted via allyl alcohol into acrolein. This reaction requires the presence of alcohol dehydrogenase. Histochemical studies have shown that this enzyme is localized in the periportal region of the liver lobule, and may explain why allyl formate solely produces a periportal necrosis. As glutathione and 1,4-dithiothreitol protect against the early biochemical changes produced by the poison, it is probable that acrolein alkylates proteins and nucleic acids.     

Production of succinate and polyhydroxyalkanoate from substrate mixture by metabolically engineered Escherichia coli

The strategic design of this study aimed at producing succinate and polyhydroxyalkanoate (PHA) from substrate mixture of glycerol/glucose and fatty acid in Escherichia coli. To accomplish this, an E. coli KNSP1 strain derived from E. coli LR1110 was constructed by deletions of ptsG, sdhA and pta genes and overexpression of phaC1 from Pseudomonas aeruginosa. Cultivation of E. coli KNSP1 showed that this strain was able to produce 21.07 g/L succinate and 0.54 g/L PHA (5.62 wt.% of cell dry weight) from glycerol and fatty acid mixture. The generated PHA composed of 58.7 mol% 3-hydroxyoctanoate (3HO) and 41.3 mol% 3-hydroxydecanoate (3HD). This strain would be useful for complete utilization of byproducts glycerol and fatty acid of biodiesel production process.     

Catabolite Inactivation in the Methylotrophic Yeast Pichia pastoris

Inactivation of the alcohol oxidase enzyme system of Pichia pastoris, during the whole-cell bioconversion of ethanol to acetaldehyde, was due to catabolite inactivation. Electron microscopy showed that methanol-grown cells contained peroxisomes but were devoid of these microbodies after the bioconversion. Acetaldehyde in the presence of O₂ was the effector of catabolite inactivation. The process was initiated by the appearance of free acetaldehyde, and was characterized by an increase in the level of cyclic AMP, that coincided with a rapid 55% drop in alcohol oxidase activity. Further enzyme inactivation, believed to be due to proteolytic degradation, then proceeded at a constant but slower rate and was complete 21 h after acetaldehyde appearance. The rate of catabolite inactivation was dependent on acetaldehyde concentration up to 0.14 mM. It was temperature dependent and occurred within 24 h at 37°C and by 6 days at 15°C but not at 3°C. Alcohol oxidase activity was psychrotolerant, with only a 17% decrease in initial specific activity over a temperature drop from 37 to 3°C. In contrast, protease activity was inhibited at temperatures below 15°C. When the bioconversion was run at 3°C, catabolite inactivation was prevented. In the presence of 3 M Tris hydrochloride buffer, 123 g of acetaldehyde per liter was produced at 3°C, compared with 58 g/liter at 30°C. By using 0.5 M Tris in a cyclic-batch procedure, 140.6 g of acetaldehyde was produced.     

Novel cellulose-based polyelectrolytes synthesized via the click reaction

Novel polyelectrolytes were prepared by conversion of 6-azido-6-deoxycellulose with acetylenedicarboxylic acid dimethyl ester and subsequent saponification. Up to 62% of the azide moieties were converted. The reaction was completed within 48 h using 2 moles of acetylenedicarboxylic acid dimethyl ester per mole of modified anhydroglucose unit. FTIR and NMR spectroscopy were applied to elucidate the molecular structure of the polymers. The polymer degradation was acceptable during this two-step reaction. The resulting biopolymer derivatives were water soluble and reduced the surface tension on water significantly. Moreover, they form ionotropic gels with multivalent metal ions.     

Evaluation and modelling the utility of SCCO2 to support efficient lipase mediated esterification

Supercritical fluids offer environmental advantages over chemical solvents, while providing enhanced separation and chemical selectivity. The use of supercritical fluids for the recovery of products from biomass and the transformation of selected molecules (to add value) was studied. Free fatty acids were bio-catalytically transformed to fatty acid esters using lipase within a supercritical fluid environment. A central composite rotatable design was used to evaluate the influence of operating conditions on the enzymatic esterification process and a response surface equation was optimized to identify the most favourable process conditions for maximum free fatty acid conversion. Based on the model equation the process conditions under which it was predicted a yield of 100% esters could be obtained were: pressure 200 bar, temperature 60 °C, ethanol concentration 2.0 M, enzyme concentration 11 wt.% and time 60 min. Experiments conducted under these conditions gave an ester yield of 94.3% (close to predicted results). The activity per unit mass of biocatalyst was found to be 1585 μmol/min/g_cat. The results support the use of supercritical fluids for process integration.     

C-N-palladacyclic-catalyzed Heck reaction in EGME/water: Rate and regioselectivity controlled by the solvents ratio

Novel as well as known C,N-palladacyclic neutral complexes were tested as precatalyst in the Heck reaction between bromobenzene and styrene under aerobic conditions. The catalytic system Pd(II) complex/K₂CO₃/EGME-H₂O (EGME = ethylene glycol monomethyl ether) showed to be highly efficient. Best performances of the catalysts were achieved by controlling the amount of water: generally a H₂O content within the 25-50% v/v range resulted in the highest conversion of the substrates into trans-stilbene. As a matter of fact, bromobenzene and styrene can be converted quantitatively using only 0.01 mol% of precatalyst with very high regioselectivity for the trans product and with a TOF of 10 000 h⁻¹. In the absence of water, all complexes were less efficient and differences in their activity were found, while such a differentiation disappeared when water was added.     

Spin labelling study of interfacial properties of egg-phosphatidylcholine liposomes as a function of cholesterol concentrations

In order to gain insight into interfacial properties of liposomes composed of egg-phosphatidylcholine (egg-PC) and dihexadecyl-phosphate (DHP) as a function of 0, 8, 15, 29, 38, 45 mol% of cholesterol, dynamic properties of two long-chain spin labels: TEMPO-stearate (2,2,6,6-tetramethylpiperidine-1-oxyl-4-yl)-octa-decanoate) and TEMPO-stearamide (2,2,6,6-tetramethylpiperidine-1-oxyl-4-yl)-octa-decanamide) were studied by CW-ESR spectroscopy. These spin labels reflect motional properties in the region of phospholipid head-groups.Two different environments of TEMPO-stearate were determined at 29, 38 and 45 mol% of cholesterol. In the newly formed domain above 29 mol%, N-O moiety of the spin label was surrounded by larger amount of bound water and experienced slower motion than in the cholesterol poor domain. The fraction of the second more hydrophilic environment of the spin label increased with cholesterol concentration. TEMPO-stearamide, a hydrogen-bond donor, reported more polar environment and slower motion than TEMPO-stearate even in the absence of cholesterol. Only one spin label environment was determined for all cholesterol concentrations. Slowing down of the TEMPO-stearamide motion was obtained even at 8 mol% of cholesterol.     

Production of extremely pure diacylglycerol from soybean oil by lipase-catalyzed glycerolysis

Research work was objectively targeted to synthesize highly pure diacylglycerol (DAG) with glycerolysis of soybean oil in a solvent medium of t-butanol. Three commercial immobilized lipases (Lipozyme RM IM, Lipozyme TL IM and Novozym 435) were screened, and Novozym 435 was the best out of three candidates. Batch reaction conditions of the enzymatic glycerolysis, the substrate mass ratio, the reaction temperature and the substrate concentration, were studied. The optimal reaction conditions were achieved as 6.23:1 mass ratio of soybean oil to glycerol, 40% (w/v) of substrate concentration in t-butanol and reaction temperature of 50 °C. A two-stage molecular distillation was employed for purification of DAG from reaction products. Scale-up was attempted based on the optimized reaction conditions, 98.7% (24 h) for the conversion rate of soybean oil, 48.5% of DAG in the glycerolysis products and 96.1% for the content of DAG in the final products were taken in account as the results.