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PubMed1998

Feasibility of measuring lipophilic or protein-bound drugs in the dermis by in vivo microdialysis after topical or systemic drug administration.

Our aim was to assess the microdialysis technique for determining in vivo drug levels of a lipophilic and a protein-bound model drug in the dermis. Forearm skin of healthy volunteers received topical 2% fusidic acid or 0.1% betamethasone-17-valerate formulations twice daily as occluded treatment on irritative dermatitis. Microdialysis sampling in the dermis after 48 h was without measurable drug. Hairless rats received maximized treatment with occluded applications of 10% fusidic acid or 4% betamethasone-17-valerate in ethanol for 72 h followed by microdialysis. Mean levels of betamethasone-17-valerate were 11-45 ng/ml; fusidic acid was not measurable. Systemic administration in clinical doses to rats was without measurable drug levels; increasing doses to 312 mg/kg of fusidic acid and 158 mg/kg of betamethasone-17-valerate yielded betamethasone-17-valerate levels of 25-44 ng/ml and fusidic acid levels of 10-90 ng/ml. This study demonstrates the challenges arising when using microdialysis for measuring in vivo-drug levels. For the drugs chosen it was necessary to administer very high systemic doses or apply a high topical drug concentration to obtain measurable drug levels in the dialysates. Drug levels were in the nanomolar range and demonstrated reproducible and dynamic monitoring of in vivo drug levels in the skin. Using microdialysis for sampling highly protein-bound or lipophilic drugs in the skin requires very sensitive analytical methods, and the sensitivity of the analysis is likely to be the limiting factor.

Administration, Oral↗
PubMed1994

Interaction between valproate and branched-chain amino acid metabolism.

Structural similarities between valproate metabolites and metabolites formed from the beta-oxidation of branched-chain amino acids (isoleucine, leucine, and valine) suggest that valproate may utilize key enzymes of branched-chain amino acid metabolism. Genetic deficiencies in these enzymes may decrease beta-oxidation of valproate and increase formation of valproate hepatotoxic metabolites. We attempted to determine if valproate interacts with branched-chain amino acid enzymes and also evaluated the effect of valproate on the urinary excretion of the straight-chain fatty acids butyrate (C4), valerate (C5), and hexanoate (C6). We collected dosage interval urine samples from three groups of 10 valproate patients: (1) valproate monotherapy, (2) valproate with carbamazepine, and (3) valproate with phenytoin. We also collected 12-hour urine samples from 10 normal volunteers who served as controls. Valproate caused a significant increase in the excretion of the deaminated acid metabolites of valine, isoleucine, and leucine. There were also significant increases in the excretion of the isoleucine metabolites 2-methylbutyrate and 2-methyl-3-OH-butyrate in the valproate patients. Valproate caused a significant increase in the excretion of all three of the straight-chain fatty acids evaluated, and valproate appears to inhibit the four types of acyl-CoA dehydrogenases involved in branched-chain amino acid and short- and medium-chain fatty acid metabolism.

Adult↗
PubMed2006

A method for measuring low-weight carboxylic acids from biosolid compost.

Concentration of low-weight carboxylic acids (LWCA) is one of the important parameters that should be taken into consideration when compost is applied as soil improver for plant cultivation, because high amounts of LWCA can be toxic to plants. The present work describes a method for analysis of LWCA in compost as a useful tool for monitoring compost quality and safety. The method was tested on compost samples of two different ages: 3 (immature) and 6 (mature) months old. Acids from compost samples were extracted at high pH, filtered, and freeze-dried. The dried sodium salts were derivatized with a sulfuric acid-methanol mixture and concentrations of 11 low-weight fatty acids (C1-C10) were analyzed using headspace gas chromatography. The material was analyzed with two analytical techniques: the external calibration method (tested on 11 LWCA) and the standard addition method (tested only on formic, acetic, propionic, butyric, and iso-butyric acids). The two techniques were compared for efficiency of acids quantification. The method allowed good separation and quantification of a wide range of individual acids with high sensitivity at low concentrations. Detection limit for propionic, butyric, caproic, caprylic, and capric acids was 1 mg kg(-1) compost; for formic, acetic, valeric, enanthoic and pelargonic acids it was 5 mg kg(-1) compost; and for iso-butyric acid it was 10 mg kg(-1) compost. Recovery rates of LWCA were higher in 3-mo-old compost (57-99%) than in 6-mo-old compost (29-45%). In comparison with the external calibration technique the standard addition technique proved to be three to four times more precise for older compost and two times for younger compost. Disadvantages of the standard addition technique are that it is more time demanding and laborious.

Calibration↗
PubMed1987

Influence of meal frequency on postprandial variations in the production and absorption of volatile fatty acids in the digestive tract of conscious pigs.

Five Large White pigs of 62.2 +/- 1.4 kg mean body weight were fitted with permanent catheters in the portal vein and carotid artery and with an electromagnetic flow probe around the portal vein to study the absorption of volatile fatty acids (VFA) by measuring the concentration of these metabolites in hourly blood samples, and by determining the portal blood flow rate for a period of 12 h after intake of a single 800-g meal (6% crude fiber) preceded by 12 or 24 h of fasting. The portal concentration of VFA mixture always highly exceeded the arterial concentration. The arterial concentrations of propionic, butyric, valeric and isovaleric acids were nearly null, accounting for an almost complete uptake of these VFA by the liver. Acetic acid also was taken up, but to a lesser extent. Total VFA absorption during 12 h was 64% higher (P less than .05) after 12 h (1,160 +/- 100 mmol/12 h) than after 24 h of preprandial fasting (740 +/- 83 mmol/12 h). It increased after the meal (P less than .05) from 82.3 +/- 7.8 mmol/h between the first and fourth hour to 107.8 +/- 7.5 mmol/h between the fifth and tenth hour when the preprandial fasting lasted 12 h; a nonsignificant increase also was found when fasting prior to the meal lasted 24 h. The composition of the VFA mixture was not modified by the length of preprandial fasting. With this type of diet there was a large predominance of acetic acid (52%) followed by propionic and butyric acids (36 and 8.5%, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Effects of various salts and pH on the stability of the nucleosome in chromatin fragments.

The stability of nucleosomes in long chromatin fragments was observed by differential scanning calorimetry over a wide range of solution conditions. The thermal denaturation of chromatin was characterized in general as three major transitions, although the process clearly is more complex. The three major transitions were (1) denaturation of the nucleosome, (2) base unstacking of DNA in the resulting denatured nucleoprotein, and (3) base unstacking of naked DNA. In very low salt concentrations (e.g., 2 mM sodium cacodylate), these three processes were essentially coincident (near 76 degrees C), but in medium salt concentrations (e.g., 100 mM NaCl) the nucleosome denaturation occurred first at about 69 degrees C and then base unstacking occurred at 85 degrees C. As [NaCl] was increased, all three processes were resolved with the observation of increasing amounts of naked DNA being melted, until at 2000 mM NaCl the calorimetric profile showed mainly the melting of DNA. The transition temperature for nucleosome denaturation decreased from 76 to 63 degrees C as the salt concentration increased from 1 to 600 mM. Destabilization of the nucleosome by increasing [NaCl] was also evident above 100 mM as a decrease in enthalpic change attributable to nucleosome denaturation. Similarly, as [NaCl] was increased above 100 mM, less and less denatured nucleoprotein was evident as more and more of the DNA melted as naked DNA. The fatty acid salts, sodium valerate and sodium caproate, destabilized the nucleosome but not the denatured nucleoprotein that resulted from the collapse of the nucleosome. In the series acetate, butyrate, valerate, caproate, it was clear that destabilization of the nucleosome increased as hydrophobicity (chain length) increased.(ABSTRACT TRUNCATED AT 250 WORDS)

Calorimetry, Differential Scanning↗
PubMed2007

Fermentation and elutriation of primary sludge: effect of SRT on process performance.

A primary sludge fermentation-elutriation pilot plant was operated using in-line and side-stream schemes. The influence of solids retention time, recirculation sludge flow-rate and solids concentration on the fermentation-elutriation process performance has been assessed in this paper. The use of high elutriation flows (12% of influent flow) improved the volatile fatty acids (VFA) concentration in the effluent stream. Suspended solids removal efficiency decreased in the primary settler when the solids retention time (SRT) was increased from 4 to 8 days. Disintegration step during hydrolysis process was pointed out as the main reason for that decrease. Maximum VFA productions were achieved at SRT between 6 and 8 days at the highest elutriation flow tested for both configurations. Propionic, butyric and valeric volatile fatty acids percentage increased when total solids sludge concentrations above 23,000mgl(-1) were used. Hydrogen accumulation, causing acetogenic bacteria inhibition, was indicated as the reason for C(3)-C(5) fatty acids accumulation.

Bioreactors↗
PubMed1991

Lipid metabolism in anaerobic ecosystems.

In anaerobic ecosystems, acyl lipids are initially hydrolyzed by microbial lipases with the release of free fatty acids. Glycerol, galactose, choline, and other non-fatty acid components released during hydrolysis are fermented to volatile fatty acids by the fermentative bacteria. Fatty acids are not degraded further in the rumen or other parts of the digestive tract but are subjected to extensive biohydrogenation especially in the rumen. However, in environments such as sediments and waste digestors, which have long retention times, both long and short chain fatty acids are beta-oxidized to acetate by a special group of bacteria, the H2-producing syntrophs. Long chain fatty acids can also be degraded by alpha-oxidation. Biotransformation of bile acids, cholesterol, and steroids by intestinal microorganisms is extensive. Many rumen bacteria have specific growth requirements for fatty acids such as n-valeric, iso-valeric, 2-methylbutyric, and iso-butyric acids. Some species have requirements for C13 to C18 straight-chain saturated or monoenoic fatty acids for growth.

Anaerobiosis↗
PubMed1989

[Microbiological and biochemical indices of changes in microbial ecology of rat large intestine under the effect of rifampicin].

Intragastric administration of rifampicin to rats in a dose of 40 mg/kg resulted in decreasing of the contents of grampositive cocci and lactobacilli and increasing of the number of gramnegative aerobic potentially pathogenic bacteria in feces of the experimental animals. It was noted that along with changes in the composition of the fecal microflora after the exposure to the antibiotic there were disorders in feces excretion of ammonia and amino acids such as alanine and glutamic acid as well as lactic, amber, butyric, valeric and alpha-ketoglutaric acids. Reduction of the changed biochemical indices was shown to be slower than that of the routine microbiological indices.

Animals↗
PubMed2005

Evaluation of sample recovery of malodorous livestock gases from air sampling bags, solid-phase microextraction fibers, Tenax TA sorbent tubes, and sampling canisters.

Odorous gases associated with livestock operations are complex mixtures of hundreds if not thousands of compounds. Research is needed to know how best to sample and analyze these compounds. The main objective of this research was to compare recoveries of a standard gas mixture of 11 odorous compounds from the Carboxen/PDMS 75-microm solid-phase microextraction fibers, polyvinyl fluoride (PVF; Tedlar), fluorinated ethylene propylene copolymer (FEP; Teflon), foil, and polyethylene terephthalate (PET; Melinex) air sampling bags, sorbent 2,b-diphenylene-oxide polymer resin (Tenax TA) tubes, and standard 6-L Stabilizer sampling canisters after sample storage for 0.5, 24, and 120 (for sorbent tubes only) hrs at room temperature. The standard gas mixture consisted of 7 volatile fatty acids (VFAs) from acetic to hexanoic, and 4 semivolatile organic compounds including p-cresol, indole, 4-ethylphenol, and 2'-aminoacetophenone with concentrations ranging from 5.1 ppb for indole to 1270 ppb for acetic acid. On average, SPME had the highest mean recovery for all 11 gases of 106.2%, and 98.3% for 0.5- and 24-hr sample storage time, respectively. This was followed by the Tenax TA sorbent tubes (94.8% and 88.3%) for 24 and 120 hr, respectively; PET bags (71.7% and 47.2%), FEP bags (75.4% and 39.4%), commercial Tedlar bags (67.6% and 22.7%), in-house-made Tedlar bags (47.3% and 37.4%), foil bags (16.4% and 4.3%), and canisters (4.2% and 0.5%), for 0.5 and 24 hr, respectively. VFAs had higher recoveries than semivolatile organic compounds for all of the bags and canisters. New FEP bags and new foil bags had the lowest and the highest amounts of chemical impurities, respectively. New commercial Tedlar bags had measurable concentrations of N,N-dimethyl acetamide and phenol. Foil bags had measurable concentrations of acetic, propionic, butyric, valeric, and hexanoic acids.

Acetophenones↗
PubMed1975

Nonequilibration of acetyl units from fatty acids and glucose in mammary glands of lactating mice.

Lipid synthesis by lactating mouse mammary gland slices from [1-carbon14] propionate, [1-carbon14] valerate, and [1-carbon-14] heptanoate, in the presence of glucose and from [uniformly labeled carbon-14] glucose in the presence of propionate, valerate, and heptanoate was studied. Regardless of the labeled substrate, almost all of the radioactivity incorporated into the total lipid fraction was in triglycerides. In experiments with labeled glucose and either the propionic. the valeric, or the heptylic acid, the carbon-14 fatty acids in triglycerides were almost exclusively composed of even-chain fatty acids. When the substrate was either [1-carbon-14] propionate, [1-carbon-14] valerate, or [1-carbon-14] heptanoate, in the presence of glucose, the pattern changed to one in which a larger proportion of the radioactivity was in the odd-chain fatty acids. These findings suggest that in mouse mammary gland, acetyl units produced via beta-oxidation of fatty acids may not equilibrate with those formed from oxidation of pyruvate which is derived from glucose.

Acetates↗
PubMed1975

Formation of volatile compounds and brown products in the model system n-hexanal-glycine.

During the boiling of a mixture of n-hexanal and glycine in aqueous medium, pH value 9, both soluble and insoluble brown colored pigments and volatile compounds were formed. The following volatile compounds were identified by gas chromatography and mass spectrometry; hexanone, decenone, undecenone, undecan-5-one, undecenedione, dodecenone, tridecanone, dodecanedione, tridecenone, dodecenedione, hexenyl hexanoate, caproic, valeric and butyric acids. The brown coloured pigments insoluble in water contained 0.57% of nitrogen. The presence of conjugated double-bonds, carbonyl and carboxyl groups was confirmed by spectral methods. The pigments were also separated by thin-layer chromatography on silica gel.

Aldehydes↗
PubMed1998

TPA and butyrate increase cell sensitivity to the vacuolating toxin of Helicobacter pylori.

The Helicobacter pylori toxin VacA induces large membrane-bound vacuolar compartments of late endosomal/lysosomal origin. Pre-treatment of cells with TPA and butyrate enhances the toxin induced vacuolisation up to 20 times, depending on the cell line, whereas other differentiating factors such as DMSO, EGF, valeric and retinoic acid have no effect. The higher toxin sensitivity induced by TPA does not result from an increased surface binding or endocytosis. The effect of TPA is apparent after several hours from addition and is inhibited by a PKC specific inhibitor. These data suggest that expression of cellular proteins, other than the toxin receptor(s), influences the vacuolating activity of VacA and may contribute to the sensitivity of different cell lines. The present findings define the most sensitive in vitro assay of the activity of VacA.

Animals↗
PubMed2002

Biomineralization of an organophosphorus pesticide, Monocrotophos, by soil bacteria.

AIMS: To study biomineralization of Monocrotophos (MCP) and identify the metabolites formed during biodegradation. METHODS AND RESULTS: Two cultures, namely Arthrobacter atrocyaneus MCM B-425 and Bacillus megaterium MCM B-423, were isolated by enrichment and adaptation culture technique from soil exposed to MCP. The isolates were able to degrade MCP to the extent of 93% and 83%, respectively, from synthetic medium containing MCP at the concentration of 1000 mg x l(-1), within 8 d, under shake culture condition at 30 degrees C. The cultures degraded MCP to carbon dioxide, ammonia and phosphates through formation of one unknown compound--Metabolite I, valeric or acetic acid and methylamine, as intermediate metabolites. The enzymes phosphatase and esterase, reported to be involved in biodegradation of organophosphorus compounds, were detected in both the organisms. CONCLUSIONS: Arthrobacter atrocyaneus MCM B-425 and B. megaterium MCM B-423 isolated from soil exposed to MCP were able to mineralize MCP to carbon dioxide, ammonia and phosphates. SIGNIFICANCE AND IMPACT OF THE STUDY: Pathway for biodegradation of MCP in plants and animals has been reported. A microbial metabolic pathway of degradation involving phosphatase and esterase enzymes has been proposed. The microbial cultures could be used for bioremediation of wastewater or soil contaminated with Monocrotophos.

Ammonia↗
PubMed1992

Influence of the method of forage conservation on feeding behaviour, intake and characteristics of reticulo-rumen content, in sheep fed ad libitum.

The influence of silage conservation methods on eating behaviour and the characteristics of reticulo-rumen contents was studied in sheep by comparing 3 forages, a silage without additive (WAS), one with additive (FAS) and hay (H), prepared from the same cut green forage. The sheep were rumen fistulated. WAS was very badly and FAS poorly preserved while the hay was dried under favourable conditions. The forages were fed ad libitum and the dry matter (DM) intake was 1,054, 1,241 and 1,469 g/day for WAS, FAS and H respectively. There was a single feeding in the morning. At the main meal, DM intake was 270, 317 and 388 g/day and ingestion rate 4.76, 4.56 and 4.16 g of DM/min for WAS, FAS and H respectively. There was slight recovery in ingestion around 16-19 h with hay and FAS but not with WAS. With hay, rumination lasted much longer than with the silages and began sooner after the end of the main meal. In contrast, overall rumination efficiency was the same for hay and FAS. There were fewer contractions of the reticulo-rumen with both WAS and hay. The amounts of reticulo-rumen contents were comparable for the 2 silages and higher for hay. The amounts of NH3 in the contents were the same with WAS and hay but greater with FAS. With WAS, contents were richer in butyric, valeric and caproic acids. The DM turnover rate of the contents was the same for FAS and hay and lower, but not significantly, for WAS. Accordingly, with silage, satiety seems to be rapidly reached but there was no evidence that organoleptic factors were involved. There were no problems with the digestion rate of silages. However, the factors limiting silage intake persisted throughout the diurnal cycle, and reticulo-rumen fill was affected. The more poorly preserved the silage the more these factors reduced the duration and volume of the meals, or both.

Animals↗
PubMed1989

Calcium uptake by bovine epididymal spermatozoa is regulated by the redox state of the mitochondrial pyridine nucleotides.

Immature caput epididymal sperm accumulate calcium from exogenous sources at a rate 2- to 4-fold greater than mature caudal sperm. Calcium accumulation by these cells, however, is maximal in the presence of lactate as external substrate. This stimulation of calcium uptake by optimum levels of lactate (0.8-1.0 mM) is about 5-fold in caput and 2-fold in caudal sperm compared to values observed with glucose as substrate. Calcium accumulation by intact sperm is almost entirely mitochondrial as evidenced by the inhibition of uptake by rotenone, antimycin, and ruthenium red. The differences in the ability of the various substrates in sustaining calcium uptake appeared to be related to their ability to generate NADH (nicotinamide adenine dinucleotide). Previous reports have documented that mitochondrial calcium accumulation in several somatic cells is regulated by the oxidation state of mitochondrial NADH. A similar situation obtains for bovine epididymal sperm since calcium uptake sustained by site III oxidation of ascorbate in the presence of tetramethyl phenylenediamine and rotenone was also stimulated by NADH-producing substrates, including lactate, and inhibited by substrates generating NAD+ (nicotinamide adenine dinucleotide, oxidized form). Further, calcium uptake by digitonin-permeabilized sperm in the presence of succinate was stimulated when NADH oxidation was inhibited by rotenone. The compounds alpha-keto butyric, valeric, and caproic acids, which generate NAD+, inhibited the maximal calcium uptake observed in the presence of succinate and rotenone, and the hydroxy acids lactate and beta-hydroxybutyrate reversed this inhibition. These results document the regulation of sperm calcium accumulation by the physiological substrate lactate, emphasize the importance of mitochondria in the accumulation of calcium by bovine epididymal sperm, and suggest that the mitochondrial location of the isozyme LDH-X in mammalian sperm may be involved in the regulation of calcium accumulation.

3-Hydroxybutyric Acid↗
PubMed1999

Ubiquity and diversity of dissimilatory (per)chlorate-reducing bacteria.

Environmental contamination with compounds containing oxyanions of chlorine, such as perchlorate or chlorate [(per)chlorate] or chlorine dioxide, has been a constantly growing problem over the last 100 years. Although the fact that microbes reduce these compounds has been recognized for more than 50 years, only six organisms which can obtain energy for growth by this metabolic process have been described. As part of a study to investigate the diversity and ubiquity of microorganisms involved in the microbial reduction of (per)chlorate, we enumerated the (per)chlorate-reducing bacteria (ClRB) in very diverse environments, including pristine and hydrocarbon-contaminated soils, aquatic sediments, paper mill waste sludges, and farm animal waste lagoons. In all of the environments tested, the acetate-oxidizing ClRB represented a significant population, whose size ranged from 2.31 x 10(3) to 2.4 x 10(6) cells per g of sample. In addition, we isolated 13 ClRB from these environments. All of these organisms could grow anaerobically by coupling complete oxidation of acetate to reduction of (per)chlorate. Chloride was the sole end product of this reductive metabolism. All of the isolates could also use oxygen as a sole electron acceptor, and most, but not all, could use nitrate. The alternative electron donors included simple volatile fatty acids, such as propionate, butyrate, or valerate, as well as simple organic acids, such as lactate or pyruvate. Oxidized-minus-reduced difference spectra of washed whole-cell suspensions of the isolates had absorbance maxima close to 425, 525, and 550 nm, which are characteristic of type c cytochromes. In addition, washed cell suspensions of all of the ClRB isolates could dismutate chlorite, an intermediate in the reductive metabolism of (per)chlorate, into chloride and molecular oxygen. Chlorite dismutation was a result of the activity of a single enzyme which in pure form had a specific activity of approximately 1,928 micromol of chlorite per mg of protein per min. Analyses of the 16S ribosomal DNA sequences of the organisms indicated that they all belonged to the alpha, beta, or gamma subclass of the Proteobacteria. Several were closely related to members of previously described genera that are not recognized for the ability to reduce (per)chlorate, such as the genera Pseudomonas and Azospirllum. However, many were not closely related to any previously described organism and represented new genera within the Proteobacteria. The results of this study significantly increase the limited number of microbial isolates that are known to be capable of dissimilatory (per)chlorate reduction and demonstrate the hitherto unrecognized phylogenetic diversity and ubiquity of the microorganisms that exhibit this type of metabolism.

Anaerobiosis↗