Elimination Of Glycerol Research Articles

Evaluation of Lipases from Wild Microbial Strains as Biocatalysts in Biodiesel Production

In this study, the evaluation of the catalytic behavior of several wild bacterial strains in the 1,3-selective ethanolysis of triglycerides with ethanol to produce a new type of biodiesel (Ecodiesel) that integrates glycerol as monoacylglycerols was carried out. The Ecodiesel production not only avoids the elimination of glycerol, which is largely generated as a by-product in the biodiesel industry, but also results in an increase in the biofuel yield. The wild microbial strain samples were obtained from several lipophilic organisms. In addition to evaluate the enzymatic extracts, the minimum grade of purification of the strains, necessary to obtain similar results to those attained with commercial lipases was studied. This purification treatment included a dialysis followed by a lyophilization process. Such extracts were directly used as biocatalysts in the transesterification reaction of sunflower oil with ethanol, attaining much better results (yield close to 100%) than those obtained with strains which were not submitted to the purification process (yields lower than 10%). Furthermore, the results here obtained are similar to those obtained with commercial lipases but were achieved under mild conditions and lower reaction time (2 h). In addition, the stability of the enzymatic extracts was corroborated by subsequent reactions, showing no loss of activity. Thus, this study brings to light that enzymatic extracts obtained by a very simple purification process can be economically competitive with the conventional biodiesel production methods.

Elimination of glycerol and replacement with alternative products in ethanol fermentation by Saccharomyces cerevisiae

Glycerol is a major by-product of ethanol fermentation by Saccharomyces cerevisiae and typically 2-3% of the sugar fermented is converted to glycerol. Replacing the NAD(+)-regenerating glycerol pathway in S. cerevisiae with alternative NADH reoxidation pathways may be useful to produce metabolites of biotechnological relevance. Under fermentative conditions yeast reoxidizes excess NADH through glycerol production which involves NADH-dependent glycerol-3-phosphate dehydrogenases (Gpd1p and Gpd2p). Deletion of these two genes limits fermentative activity under anaerobic conditions due to accumulation of NADH. We investigated the possibility of converting this excess NADH to NAD(+) by transforming a double mutant (gpd1∆gpd2∆) with alternative oxidoreductase genes that might restore the redox balance and produce either sorbitol or propane-1,2-diol. All of the modifications improved fermentative ability and/or growth of the double mutant strain in a self-generated anaerobic high sugar medium. However, these strain properties were not restored to the level of the parental wild-type strain. The results indicate an apparent partial NAD(+) regeneration ability and formation of significant amounts of the commodity chemicals like sorbitol or propane-1,2-diol. The ethanol yields were maintained between 46 and 48% of the sugar mixture. Other factors apart from the maintenance of the redox balance appeared to influence the growth and production of the alternative products by the genetically manipulated strains.

Modulation of activity of the adipocyte aquaglyceroporin channel by plant extracts

The plasma membrane protein, aquaglyceroporin-7 (AQP7) is exclusively expressed in adipocytes and appears to be a channel for glycerol entry and exit. It is possible that by facilitating the opening of these channels, the loss of intracellular glycerol could be encouraged and thus reduce the size of the lipid reservoir. Human preadipocytes and mouse 3T3-L1 preadipocytes were induced to develop an adipocytic phenotype by culture in a semi-defined medium. After 7 days, the expression of AQP7 message had increased by 37-fold, a level which could be further up-regulated by troglitazone or retinoic acid or down-regulated by insulin. The mature adipocytes also expressed immunoreactive aquaporin (AQP) channel protein as assessed by immunocytochemistry and Western blot. The addition of adrenaline to the culture medium stimulated the release of glycerol (blockable by HgCl(2)). Plant extracts, with potential anti-cellulite properties, were tested for their effect on glycerol elimination. These included wild yam root (Dioscorea opposita), cocoa bean (Theobroma cacao), horse chestnut tree (Aesculus hippocastanum) seed and bark and tomato (Solanum lycopersicum). Of these, D. opposita appeared to induce a dose-dependent glycerol release. The results show that our assay can help to identify modulators of AQP7 channel expression and activation in adipocytes.

Internal residue loss produced by rearrangement of a novel cationic glycosphingolipid, glyceroplasmalopsychosine, in collision-induced dissociation.

A novel plasmal conjugate of galactosylsphingosine (psychosine), Gro1(3)-O-plasmal-O-6Galbeta-sphingosine (glyceroplasmalopsychosine), was analyzed by electrospray ionization and liquid secondary ion mass spectrometry with low- or high-energy collision-induced dissociation (CID). In the product ion spectra of the [M + H](+) ions, [M + H - glycerol](+) ions arising from the loss of a glycerol were predominant. Unexpectedly, CID of the [M + H - glycerol](+) ion produced an outstanding ion, [(M + H - glycerol) - Hex](+), which required the loss of the galactose from inside the molecule. This ion was greatly reduced in the spectra of N,N-dimethyl derivatives, indicating that the [(M + H - glycerol) - Hex](+) ion is formed from an intramolecular rearrangement with migration of the plasmal residue to the free amino group of sphingosine. It would be expected that the rearrangement occurs simultaneously with the elimination of glycerol or a rearranged [M + H](+) ion leads to the elimination of glycerol, to form a Schiff base-type [M + H - glycerol](+) ion, from which the terminal galactose could be removed by the normal mechanism of glycosidic cleavage. On the other hand, the [M + Na - glycerol](+) ion derived from the sodiated molecule did not produce an ion corresponding to the rearrangement reaction, possibly owing to a higher stability of the sodiated ions against conformational changes.

Unexpected Fragmentation of a Novel Cationic Glycosphingolipid, Glyceroplasmalopsychosine, by Electrospray Ionization and Low Energy Multistage Mass Spectrometry.

Glyceroplasmalopsychosine, Gro1(3)-O-plasmal-O-6Galβ-sphingosine, is a unique plasmal conjugate of galactosylsphingosine (psychosine) that has an O-acetal conjugate of plasmal to the hydroxyl groups of both glycerol and galactosyl residues. Electrospray ionization ion trap mass spectrometry was applied to the structural analysis of this unique compound. In the MS/MS (MS2) spectrum produced by low energy collision-induced dissociation (CID) of the [M+H]+ ion, elimination of glycerol was predominant. MS/MS/MS (MS3) of the [M+H-glycerol]+ ion yielded an outstanding anomalous ion which required the loss of a hexose in the middle of the molecule with the plasmal residue retained on the psyhchosine side. This unexpected fragmentation by MS3 could be attributed to a rearrangement of the plasmal residue to the sphingosine residue, allowing elimination of the galactose by subsequent MS3 after CID.

A point mutation in the glycerol kinase gene associated with a deletion in the dystrophin gene in a familial X-linked muscular dystrophy: non-contiguous gene syndrome involving Becker muscular dystrophy and glycerol kinase loci

We report a family with an X-linked recessive muscular dystrophy characterised by exercise-induced myalgia, recurrent pigmenturia and mild proximal muscle involvement. Immunocytochemical and immunoblotting analysis in muscle, using the antibody directed against the rod domain of dystrophin, revealed a loss of immunoreactivity, but the immunolabelling using the antibodies directed against the COOH and NH 2 domains of dystrophin were almost normal. The immunoreactions for α-sarcoglycan, γ-sarcoglycan and β-dystroglycan were normal. In the five male patients of this family with increased serum creatine kinase levels (from ×8 to ×50), mass spectrometry screening of the urine revealed a large increase in glycerol elimination which was quantified by enzymatic assay (from ×14 to ×39). An in-frame deletion of the dystrophin gene (exons 13–29) was found in the same five males and in three carrier females. All the deleted chromosomes also carried a missense mutation at nucleotide 947 of the Xp glycerol kinase (GK) gene resulting in a Thr to Met substitution at codon 278. These findings indicate that the two mutations cosegregate on the same chromosome in this family. This is the first reported case of two physically independent mutations, within the DMD and GK genes, which are contiguous but several hundred kilobases apart.

Complexation of transition-metal ions, SnII, PbII and Al III with nucleobase-substituted polyethers and dissociation of adduct ions studied by fast atom bombardment mass spectrometry

Metal-ion-complexed dinucleotide analogues were studied by FAB mass spectrometry using tri- and bi-valent transition-metal ions (Cr to Zn), Sn II , Pb II and Al III . Pairs of thymine nucleobases linked by (CH 2 ) 2 - [O(CH 2 ) 2 ] n O(CH 2 ) 2 chains (n = 1 1, 2 2, 3 3 or 4 4) and of adenine nucleobases (n = 1 5 or 4 6) co-ordinate to metal ions and form significant [M + M - H] + and [M + MCl] + ions. In addition, many transition-metal ions also give [M + M 2 X - 2H] (X = Cl or NO 3 ) ions but Cr III produces [M + CrCl 2 ] + ions. The effect of the spacer chain length is reflected in the ease of formation of [M + M - H] + and [M + MX] + ions. Fragmentation of [M + MX] + and [M + M 2 X - 2H] ions give [M + M - H] + ions, suggesting that metal chelation through the nucleobase is more favoured than through the polyether. Complexes of compounds 1–6 with Al III –glycerol give abundant [M + 117] + complex ions (117 = Al + glycerol - 2H). The intensities of these ions decrease with increasing number of ethylene oxide units from 1–4. The same trend is also observed in 5 and 6. Bimetallic ions with an unipositive charge are also produced from 1–4. Dissociation of adduct ions labelled with deuterium confirms the substitution of amide protons with aluminium(III) in the elimination of glycerol.

Temporary reversal of serum to cerebrospinal fluid glycerol concentration gradient after intravenous infusion of glycerol.

Glycerol 50 g infused i.v. over 2 to 6 h is widely used to treat cerebral oedema in patients with acute stroke. Its transit through the blood-cerebrospinal fluid barrier in subjects with uninflamed meninges has now been examined. In 7 patients with an external ventriculostomy for occlusive hydrocephalus, each of whom was given 500 ml of a 10% solution IV over 4 h, serum and CSF were repeatedly sampled during and after the infusion and glycerol was measured enzymatically. The highest serum glycerol level of 191-923 mg/l was observed at the end of the infusion. The maximum CSF glycerol of 18.7-110.8 mg/l was attained 0-1 h after the end of the infusion. Elimination both from serum and CSF approximated a single-exponential decay; the elimination half-life from serum was 0.29-0.56 h compared to 1.03-3.68 h from CSF. In six of the seven cases there was a temporary reversal of the serum/CSF concentration gradient during glycerol elimination. The ratios of the AUCs of CSF and serum, which describe the overall penetration of glycerol into CSF, ranged from 0.09-0.31. In conclusion, the serum level of glycerol produced by giving 50 g IV glycerol over 4 h may not be sufficiently high reliably dehydrate to brain tissue in many patients, and the slow elimination of glycerol from the CSF may be related to the so-called rebound phenomenon.

The kinetics of the splanchnic elimination of ethanol and glycerol.

Abstract: The kinetics of the splanchnic ethanol‐ and glycerol‐elimination were studied in fasting cats by simultaneous estimation of the splanchnic elimination rate and the average splanchnic concentration of these substances. Ethanol elimination followed Michaelis kinetics with a maximal elimination rate of 27 μmol/kg/min. and apparent in vivo Km of 0.25 mM. In contrast, the kinetic constants in glycerol elimination could not be determined, probably due to a splanchnic production of glycerol and/or a diffusion barrier for glycerol from blood to hepatic cytosol.

Glycerol and free fatty acid turnover in leukaemia and hyperthyroidism.

Blood glyccrol and plasma FFA were within the normal range in leukaemic patients with hypermetabolism. The fractional turnover rates of glycerol were increased by about 55% and the turnover rates by about 35%. The leukocytes are thought to be responsible for the raised elimination of glyce rol in the leukaemic state. Results suggest that fat mobilization in leukaemics is less than in hyperthyroids on comparable basal metabolic rate. Enhanced protein metabolism accounts for the remaining part of the energy expendi ture during basal conditions in leukaemia.

Glycerol uptake in leukocytes and thrombocytes.

The glycerol disappearance was studied in whole blood and in leukocyte and thrombocyte suspensions incubated at 37°. The glycerol uptake rate was 0.38 μmoles per hour in 109 leukocytes and 3.9 in 1011 thrombocytes at a glycerol concentration of 70 μM. The uptake rate was reduced in leukaemic leukocytes and increased in leukocytes from patients with myelo fibrosis and infectious leukocytosis. In the thrombocytes also it varied with disease. The results indicate that leukocytes and thrombocytes are respon sible for a significant part of the glycerol elimination in man.

Glycerol turnover in hyperthyroidism

The fasting blood glycerol concentrations, determined from capillary samples, averaged 0.048 m M for euthyroid subjects and 0.136 m M for hyperthyroid patients, when studied in supine position. The mean value for the euthyroid significantly decreased as compared to that obtained with subjects sitting. This fact strongly emphasizes the necessity of strictly standardized conditions before blood collection when studying the variation in glycerol concentration in man. The mean glycerol turnover rate was i.04μmoles/min/kg for the euthyroids and 3.44 for the hyperthyroids. A highly significant correlation was obtained between the turnover rate and the blood glycerol concentration in the two groups. The mean fractional turnover rate of the hyper-thyroid group was only increased by 15%, demonstrating very slight influence of the hyperthyroid state on the glycerol elimination. The determination of glycerol enabled euthyroid and hyperthyroid subjects to be distinguished with only a slight degree of overlap. By determining the glycerol turnover rate, overlap was entirely avoided.

Elimination of Glycerol as a Measure of the Hepatic Blood Flow in the Cat

AbstractThe metabolism of glycerol has been studied in eleven cats anesthetized with Nembutal. The elimination rate after a single injection of glycerol was found to be constant down to arterial concentrations of 100—150 mg/l blood. The elimination rate averaged 257.5 mg/kg/h or 10.35 mg/min. Below arterial concentrations of 100—150 mg/l blood the extraction in the liver seemed to be complete. By continuous infusion of small amounts of glycerol it was demonstrated by changing the amount infused per minute at least once that the liver blood flow could be calculated without catheterization of the liver. The mean value of the flow calculated this way was 57.2 ml/kg/min. An extrahepatic elimination of glycerol was demonstrated in the eviscerated animal, averaging 2.38 mg/min. Apparently the extrahepatic elimination took place in the kidneys.