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PubMed1994

Insulin stimulates thyroid hormone receptor alpha gene expression in cultured bovine aortic endothelial cells.

Thyroid hormones and insulin regulate numerous cell processes and potentially interact through the transcriptional regulation of key genes. For instance, thyroid hormones stimulate the transcription of the fatty acid synthase and malic enzyme genes in chick embryonic hepatocytes, while insulin amplifies these effects. It is possible that insulin augments these actions of thyroid hormone by stimulating production of the thyroid hormone nuclear receptor (TR). In these studies, we examined the regulation of TR production/gene expression by insulin in bovine aortic endothelial cells (BAEC). We demonstrate that insulin significantly stimulates the gene expression of the TR alpha receptor, from BAEC. Insulin causes a maximal threefold induction above control TR alpha steady state mRNA levels in time and dose-related fashion in these cells. The increased mRNA mainly resulted from a twofold increase in transcription, as determined by nuclear run on. Insulin also increases thyroid receptor number and thyroid hormone binding, determined by Scatchard analysis of competitive inhibition binding studies. An established observation is that insulin can synergistically augment thyroid hormone-induced transcriptional activation of several important genes. It has also been previously determined that thyroid hormone action correlates closely to TR nuclear receptor number. Therefore, our studies, which show that insulin stimulates TR alpha production, suggests a potential mechanism whereby insulin can augment thyroid hormone transcriptional action.

Animals↗

PubMed2005

Comparison of highly-fluorinated chloroformates as direct aqueous sample derivatizing agents for hydrophilic analytes and drinking-water disinfection by-products.

Four highly-fluorinated alkyl and aryl chloroformates, including 2,2,3,3,4,4,5,5-octafluoro-1-pentyl chloroformate (OFPCF), 2,3,4,5,6-pentafluorobenzyl chloroformate (PFBCF), 3,3,4,4,5,5,6,6,7,7,8,8,8-tridecafluoro-1-octyl chloroformate (TDFOCF), and 2-(2,3,4,5,6-pentafluorophenoxy)-ethyl chloroformate (PFPECF), were synthesized and tested as reagents for the direct water derivatization of polar and hydrophilic analytes. The goal of this research was to develop an optimal derivatizing agent to aid in the identification of highly polar ozonation drinking water disinfection by-products (DBPs) that are believed to be missed with current analytical procedures. The chemical properties (reactivity, selectivity, derivatization products, and their chromatographic and spectral features) for the four chloroformates were investigated using a set of highly polar standard analytes, including malic and tartaric acids, hydroxylamine, valine, 2-aminoethanol, resorcinol, 1,3,5-trihydroxybenzene, and 2,4-dihydroxybenzoic acid. Upon derivatization, the analytes were extracted from the aqueous solvent and analyzed by gas chromatography (GC)-mass spectrometry (MS) in the electron capture negative ionization (ECNI) mode. Positive chemical ionization (PCI)-MS was used for confirmation of molecular ions that were weak or absent in ECNI mass spectra. Of the four derivatizing reagents tested, OFPCF showed the best performance, with good reaction efficiency, good chromatographic and spectroscopic properties, low detection limits (10-100 fmol), and a linear response more than two orders of magnitude. Further, the entire procedure from raw aqueous sample to ready-to-inject hexane solutions of the derivatives requires less than 10 min. PFBCF showed ideal applicability for derivatizing aminoalcohols and aminoacids. The two chloroformates with the highest intrinsic stability (TDFOCF and PFPECF) failed to derivatize some of the analytes. Finally, the OFPCF derivatizing agent was tested with simulated ozonated drinking water (aqueous fulvic acid treated with ozone), and three highly polar reaction by-products were determined.

Journal Article↗

PubMed2002

Differential role of PPAR gamma in the regulation of UCP-1 and adipogenesis by TNF-alpha in brown adipocytes.

Extracellular regulated kinases (ERKs) mediate the inhibitory effect of tumor necrosis factor alpha (TNF-alpha) on uncoupling protein-1 (UCP-1), but not on lipid accumulation. TNF-alpha-induced ERK-dependent peroxisome proliferator activator receptor gamma (PPAR gamma) phosphorylation could be responsible for UCP-1 downregulation. Thus, the negative effect of TNF-alpha on UCP-1 mRNA expression at 4-5 h, under basal conditions or in cells treated with the PPAR gamma agonist, rosiglitazone, was reversed by the MEK1 inhibitor PD98059. In contrast, fatty acid synthase and malic enzyme mRNA downregulation was not prevented. Moreover, rosiglitazone has no positive effect on adipogenic gene expression or lipid accumulation. Therefore, there is a differential regulation of thermogenic and adipogenic differentiation by PPAR gamma, which might account for the differences in the TNF-alpha regulation through ERKs.

3T3 Cells↗

PubMed2002

Moist heat treatment on physicochemical change of chitosan salt films.

Chitosan salt films were prepared by casting method using acetic, citric, formic, glycolic, lactic, malic and propionic acids as solubilizers. The films were then exposed to moist heat at 60 degrees C and 75% relative humidity for several time intervals. The influence of moist heat treatment on their physicochemical characteristics was investigated. All freshly prepared films were soluble in deionized water and HCl buffer solution. Chitosan citrate film also dissolved in phosphate buffer solution. After treatment, the percentage of water sorption and dissolution of chitosan films in three media were gradually decreased. Longer alkyl group and less carboxyl and hydroxyl groups in the molecule of organic acid resulted in lower percentage of water sorption and dissolution of treated films. The FT-IR spectra revealed that there was amide formation between chitosan and organic acids after treatment especially in chitosan acetate and propionate films. However, the absorption peaks of ammonium ['NH3+] and free carboxylate groups were still remained in treated chitosan citrate and malate films. Change in the degree of crystallinity from powder X-ray diffractogram and thermal characteristic from DSC thermogram were also related to the water sorption and dissolution of films.

Chemical Phenomena↗

PubMed2001

Differential effects of diets that provide different lipid sources on hepatic lipogenic activities in rats under ad libitum or restricted feeding.

This work was designed to study the effect of different lipid sources on hepatic lipogenic enzyme activity in rats fed ad libitum or energy-controlled diets. Male Wistar rats were fed diets containing 40% of energy as fat (olive oil, sunflower oil, palm oil, or beef tallow) for 4 wk. In experiment 1 rats had free access to food, and in experiment 2 rats were fed a controlled amount of food. In both experiments, rats fed the olive oil diets had higher activities of glucose-6-phosphate dehydrogenase, malic enzyme, fatty acid synthase, and acetyl-CoA carboxylase (P < 0.05) than rats fed the other fats. It is unlikely that this effect could be attributed to the stimulation by insulin or triiodothyronine because serum values did not differ among the groups. Enzymatic activities were positively and significantly correlated with liver triacylglycerol content, but not with serum triacylglycerol levels. No interaction between lipid source and feeding protocol was found. Oleic acid and components in olive oil other than fatty acids, such as phytosterols, may account for the effects of dietary fat on lipogenic enzyme activity.

Acetyl-CoA Carboxylase↗

PubMed2005

Extending the lifetime of the running electrolyte in capillary electrophoresis by using additional compartments for external electrolysis.

The use of two additional reservoirs to accommodate the electrodes of the power source is proposed to improve the stability of the running electrolyte in capillary electrophoresis. The basic idea is to use salt bridges to connect those reservoirs to the ones containing the capillary ends. Although simple, there are several issues that can be considered in the design and implementation of such system in order to prevent undesired transference of material between the electrolysis and the main reservoirs. The use of a sealed electrolysis reservoir without a gas phase, the use of materials that ensure volume stability, and the use of bridges as long as possible are three basic directions. A compromise is involved in the dimensions of the sectional area of the bridge, because a small area diminishes the amount of a species transferred by diffusion but leads to an undesirable increase of the electrical field during the electrophoretic running. Thus, a bridge composed of a main wide-bore tube connected to a small-bore capillary seems to give the best performance for practical use. A simple electrolysis-separated system was adapted to a preexisting capillary electrophoresis system, and its performance was evaluated with a mixture of tartaric, malic, and succinic acids that was separated in sodium benzoate solution (pH 5.5) using the original equipment and the modified one. Due to the water electrolysis and the small buffering capacity of the electrolyte, there was a significant pH change and consequently changes in the effective mobilities of the analytes and loss of resolution after a few runs using the original equipment. Using the electrolysis-separated system, no significant change in the migration time and resolution was observed even after 15 runs. Besides the freedom to prepare running electrolytes with electroactive species or unbuffered solution, high throughput and the use of small reservoirs, such as the ones used in microfluidic devices, are the main advantages of the system.

Journal Article↗

PubMed1988

Two randomized controlled trials of virucidal nasal tissues in the prevention of natural upper respiratory infections.

Two six-month randomized, controlled double-blind trials of the efficacy of virucidal nasal tissues in the prevention of natural colds were conducted in Charlottesville, Virginia, between 1983 and 1986. Tissues impregnated with malic and citric acids and sodium lauryl sulfate were used in both trials. Placebo tissues contained saccharin in the first trial and a mixture including succinic acid in the second. A total of 186 families were evaluable after completion of trial I, and 98 families were evaluable in trial II. The antiviral tissues were associated with 14 and 5% relative reductions in the overall rate of colds in the first and second trials, respectively. In the first study, this appeared to be due to an appropriate fall in secondary illnesses with a relative reduction in the ratio of secondary to primary illnesses of 32%. In trial II, however, the small and statistically insignificant reduction was primarily due to a drop in the rate of primary illness (which cannot be attributed to tissue efficacy), and the ratio of secondary to primary illness was actually 5% higher in the group with active tissues than in the placebo group. We conclude that when rigorously used in a study protocol, virucidal tissues may offer a modest reduction of secondary colds in the home, but for reasons currently unknown, do not have a major effect on the overall rate of colds.

Antiviral Agents↗

PubMed1975

Iron deficiency and the structure and physiology of maize chloroplasts.

The ultrastructure of mesophyll chloroplasts of maize (Zea mays L.) was more severely affected by iron deficiency that induced mild chlorosis than was the ultrastructure of bundle sheath plastids. Ferredoxin and ribulose diphosphate carboxylase levels were severely decreased by iron deficiency. Malic enzyme was less affected, and phosphoenolpyruvate carboxylase activity remained high even under severe iron deficiency. Iron deficient leaves fixed carbon into malic and aspartic acids but the rate of entrance of carbon into the sugar phosphates and sucrose was greatly reduced compared to the control. Chlorophyll a/b ratios ranged from low values of less than 2 in severely iron deficient leaves to high values exceeding 4 in leaves showing little iron deficiency.

Journal Article↗

PubMed1986

Function of Rhizodermal Transfer Cells in the Fe Stress Response Mechanism of Capsicum annuum L.

A variety of red pepper (Capsicum annuum L., cv Yaglik) responds to Fe deficiency stress with simultaneously enhanced H(+) extrusion, reduction of ferric ions and synthesis of malic and citric acid in a swollen subapical root zone densely covered with root hairs. It is demonstrated that these stress responses temporally coincide with the development of rhizodermal and hypodermal transfer cells in this root zone. During stress response the transfer cells show a marked autofluorescence which could arise from endogenous iron chelators of the phenolic acid type. The presence of organelle-rich cytoplasm which often exhibits rotational cytoplasmic streaming points to high physiological activity and makes these cells, with their increased plasmalemma surface, particularly well suited for the entire stress response mechanism. Since Fe stress-induced acidification is diminished by vanadate and erythrosin B, both specific inhibitors of plasmalemma ATPases, it seems reasonable to suppose that H(+) pumping from transfer cells is activated by an ATPase located in their plasmamembrane. H(+) extrusion is also shown to be inhibited by abscisic acid. Raised phosphoenolpyruvate carboxylase activity and simultaneous accumulation of malate in the swollen root zone point to the action of a pH stat preventing a detrimental rise in cytoplasmic pH of transfer cells during enhanced H(+) extrusion. The simultaneous increase in citric acid concentration favors chelation of iron at the site of its uptake and thus ensures long distance transport to the areas of metabolic demand. A direct link between citrate accumulation and ferric ion reduction as proposed in recent literature further supports the crucial role of transfer cells in the response to Fe deficiency stress.

Journal Article↗

PubMed2006

Transcriptional regulation by insulin: from the receptor to the gene.

Insulin, after binding to its receptor, regulates many cellular processes and the expression of several genes. For a subset of genes, insulin exerts a negative effect on transcription; for others, the effect is positive. Insulin controls gene transcription by modifying the binding of transcription factors on insulin-response elements or by regulating their transcriptional activities. Different insulin-signaling cascades have been characterized as mediating the insulin effect on gene transcription. In this review, we analyze recent data on the molecular mechanisms, mostly in the liver, through which insulin exerts its effect. We first focus on the key transcription factors (viz. Foxo, sterol-response-element-binding protein family (SREBP), and Sp1) involved in the regulation of gene transcription by insulin. We then present current information on the way insulin downregulates and upregulates gene transcription, using as examples of downregulation phosphoenolpyruvate carboxykinase (PEPCK) and insulin-like growth factor binding protein 1 (IGFBP-1) genes and of upregulation the fatty acid synthase and malic enzyme genes. The last part of the paper focuses on the signaling cascades activated by insulin in the liver, leading to the modulation of gene transcription.

Gene Expression Regulation↗

PubMedDate not supplied

Messenger RNA levels and transcription rates of hepatic lipogenesis genes in genetically lean and fat chickens.

Levels of body fat content in commercial meat chickens have prompted research in order to control the development of this trait. Based on experimentally selected divergent lean and fat lines, many studies have shown that liver metabolism has a major role in the fatness variability. In order to identify which genes are involved in this variability, we investigated the expression of several genes implicated in the hepatic lipid metabolism. The studied genes code for enzymes of fatty acid synthesis [ATP citrate-lyase (ACL), acetyl-CoA carboxylase (ACC), fatty acid synthase (FAS), malic enzyme (ME), stearoyl-CoA desaturase (SCD1)], for an apolipoprotein [apolipoprotein A1 (APOA1)], and for the CCAAT/enhancer binding protein alpha (C/EBPalpha), which is a transcription factor implied in the regulation of several genes of lipid metabolism. The results show that the fat-line chickens display significantly higher hepatic transcription rates and mRNA levels than the lean-line chickens for the ACL, ME and APOA1 genes. This suggests that these genes could be responsible for the phenotypic fatness variability.

Journal Article↗

PubMed2006

Low birth weight is associated with enlarged muscle fiber area and impaired meat tenderness of the longissimus muscle in pigs.

The objective of this study was to determine the relationships between birth-weight-associated modifications in histological or chemical muscle characteristics and meat quality traits in pigs. At 68 d of age, Pietrain x (Large White x Landrace) female littermates were allocated into 2 groups on the basis of low birth weight (LW = 1.05 +/- 0.04 kg; n = 15) or high birth weight (HW = 1.89 +/- 0.02 kg; n = 15). Pigs were reared in individual pens with free access to a standard diet up to slaughter at approximately 112 kg of BW. During the growing-finishing period, LW and HW pigs had a similar daily feed consumption, whereas G:F was lower (P = 0.009) for LW pigs than for HW littermates. At final BW, LW pigs were 12 d older (P < 0.001) than HW littermates. Estimated lean meat content, relative proportions of loin and ham in the carcass, and weights of LM and semitendinosus muscle (SM) were decreased (P < 0.05) in LW pigs compared with HW pigs. Conversely, the LW pigs exhibited a fatter carcass, greater activity levels of fatty acid synthase and malic enzyme in backfat (n = 15 per group), and enlarged subcutaneous adipocytes (n = 8 per group) compared with the HW pigs. Similarly, lipid content was increased by 25% (P = 0.009), and mean adipocyte diameter was 12% greater (P = 0.008) in the SM from LW pigs compared with that from HW pigs, whereas lipid content did not vary in the LM of either group. Mean myofiber cross-sectional areas were 14% greater in the LM (P = 0.045) and the SM (P = 0.062) of LW pigs than of HW pigs. Conversely, the total number of myofibers was less (P = 0.003) in the SM of LW vs. HW pigs. There were no differences between groups for glycolytic potential at slaughter and rate and extent of postmortem pH decline in both muscles, as well as for LM drip losses. A trained sensory test panel judged the roast loin meat to be less tender (P = 0.002) in LW pigs relative to HW pigs. Scores for juiciness, flavor, flouriness, and fibrousness of meat did not differ between groups. Overall, negative but somewhat low correlation coefficients were found between LM tenderness score and ultimate pH (r = -0.36; P = 0.06) and between LM tenderness and mean cross-sectional area of myofibers (r = -0.34; P = 0.07). This study demonstrates a lower tenderness of meat from pigs that had a LW, partly as a result of their enlarged myofibers at market weight.

Animals↗

PubMed1983

Wheat pasture poisoning. I. An evaluation of cereal pastures as related to tetany in beef cows.

Forage samples were obtained from three wheat and rye mixed pastures (replications) from December 1979, to April 1980 (140 d), while they were being grazed by mature cows at the Southwestern Livestock and Forage Research Station, El Reno, Oklahoma. These samples were taken three times during December, twice monthly in January and February and weekly in March and April. Samples were analyzed for dry matter, ash, N, K, Ca, Mg, P, ash alkalinity, aconitic, malic and citric acids, total lipids, NO3-N, Na and total nonstructural carbohydrates (TNC). In vitro digestibility was also determined. The N:TNC ratio, estimated plasma Mg levels and K:(Ca + Mg) ratio (tetany ratio) were calculated. On d 105 (March 19), 16% (five of 32 head) of the cows developed tetany. At tetany, the forage K, protein, digestibility, ash alkalinity, aconitic acid and total lipids increased suddenly and markedly. Forage dry matter, TNC and ash decreased. Forage Ca and Mg were slightly below or equal to the animals' requirements and remained relatively constant during the period of tetany. Forage P increased about 2 wk before tetany and decreased after tetany occurred. The levels of forage NO3-N and Na did not appear to be related to the incidence of tetany. The N:TNC ratios exceeded .4 immediately before tetany and two sampling periods thereafter. The estimated plasma Mg levels were lowest at tetany and shortly thereafter and tetany ratio exceeded 2.2 during most of the study, with peaks of 3.2 in December, at tetany and in early April.

Animals↗

PubMed1994

Monostearoylglycerol-starch complex: its digestibility and effects on glycemic and lipogenic responses.

We examined whether a modification of a starch into an alpha-amylase resistant form can lead to a reduction of postprandial glucose and insulin responses, and consequently to a change of lipid metabolism in liver and adipose tissue. For this purpose, a processed starch was prepared using a cornstarch (70% amylose and 30% amylopectin) and monoacylglycerol (monostearate; MS), forming monostearate-starch complex (MS-treated cornstarch). When we determined in vitro hydrolysis of MS-treated cornstarch using alpha-amylase and intestinal microvillar alpha-glucosidases, the glucose production rate of the MS-treated cornstarch was slower than the non-treated cornstarch. Measurement of a transmural potential difference (delta PD) evoked by the MS-treated cornstarch in everted rat jejunum showed that the absorption rate of glucose released from the MS-treated cornstarch was also remarkably slower than that from the non-treated cornstarch. The postprandial plasma insulin response to the MS-treated cornstarch was reduced, although plasma glucose response was unchanged. In a feeding study, two groups of five or six male Wistar-strain rats were fed defined diets containing 61.1% MS-treated cornstarch or 58.2% non-treated cornstarch ad libitum for 14 days. Food intakes during the period were similar between the two groups. Feeding the MS-treated cornstarch resulted in a significantly lower maltase activity in upper jejunum than did the non-treated cornstarch feeding. The activities of lipogenic enzymes--fatty acid synthetase (FAS), malic enzyme (ME), and glucose-6-phosphate dehydrogenase (G-6-PDH)--significantly decreased in epididymal adipose tissue of rats fed the MS-treated cornstarch. In the liver, FAS activity was lower in the MS-treated cornstarch group. The results indicated that MS-treated cornstarch was digested less rapidly, and lowered blood insulin response, consequently leading to a declined lipogenesis of adipose tissue and liver. This study suggests that the rate of intestinal hydrolysis of starch is an important determinant of metabolic responses such as glycemic and lipogenic responses to diets.

Adipose Tissue↗

PubMed1985

Effect of dietary fat and environment on lipogenesis by large white breeder turkeys.

An experiment was conducted to evaluate the effects of either a low-fat (8% of total calories) or high-fat (47% of total calories) diet on Large White breeder females maintained at a constant (21 C) or a cyclic temperature regimen (12 to 27 C). Each diet was fed to 32 hens individually caged in the cyclic condition and 46 in the constant condition. Energy consumption, reproductive performance, and in vitro lipogenesis were monitored for hens subjected to both temperature and dietary treatments. Hens consumed more (P less than .05) dietary energy when maintained at a constant temperature or when fed the low-fat diet. Hens also produced more (P less than .05) eggs when maintained under cyclic temperature conditions, although this observation was tempered by the fact that fewer hens were broody. More (P less than .05) of the fertile eggs from hens held at the cyclic temperature resulted in live poults. The high-fat diet decreased (P less than .05) in vitro lipogenesis in hens and may also decrease the maintenance energy requirement by providing a preformed fatty acid pool for egg lipids. The high-fat also decreased (P less than .05) liver lipid and the activities of fatty acid synthetase and malic enzyme.

Animals↗

PubMedDate not supplied

Morphogenesis and pathogenesis of chronic lung diseases. XI. Histoenzymatic study of the experimentally developing lung granulomas.

The development of lung granulomas induced in rabbits by an intravenous injection of the complete Freund adjuvant containing dog pulmonary proteins was followed by histologic and histoenzymatic methods. By comparison with normal rabbit lungs. the developing structures of the induced lung granulomas showed an increase of all tested enzymes at 15 days, and a secondary intensity decrease of the activity of malic dehydrogenase, of acid and alkaline phosphatases, and of alanyl-aminopeptidase at 30 days. Only the hydroxyproline-epimerase showed at these intervals an increasing activity in the lung granulomas. The developed reticulinic fibrils appeared to have a highly intense alkaline phosphatase activity.

Animals↗

PubMed1985

Purification and characterization of Ricinus communis invertase.

An invertase from Ricinus communis leaves was purified 4,400-fold. The preparation was homogeneous by criteria of gel electrophoresis, gel permeation, adsorption, and ionic exchange chromatography. One optimum pH at 3.5 was observed with crude invertase; however, purified preparations showed two optima, at pH 3.5 and 5.5. Addition of bovine serum albumin restored one maximum at pH 3.5 and elicited a 30% activation of the invertase. The effect was caused by many other proteins and by heparin, dextran sulfate, and polyvinylpyrrolidone. Fructose, fructose 1,6-diphosphate, maleic, trans-aconitic, malic, and ascorbic acids were simple competitive inhibitors of the purified enzyme. Glucose was a noncompetitive inhibitor. The activation by proteins suppressed these inhibitory effects. The minimum concentration of activator necessary to reach the maximal activation or "point of optimal activation" was always reached at a concentration of 1 X 10(-6) M, independently of the nature of the activator, when 8.6 X 10(-12) mol of enzyme were used. Apparent molecular weight determinations of the enzyme in the presence and absence of activator and molecular weight determinations based on determinations of the point of optimal activation suggested that the purified enzyme is a heptamer (Mr of 77,900, Stokes radius 32 A, frictional ration f/fo 1.1, partial specific volume 0.749 ml/g) and that the activated form is a trimer consisting of two enzyme subunits and one activator molecule. The activation was lost by dilution of the trimer. The enzyme subunit, as isolated by gel filtration in the presence of sodium dodecyl sulfate (Mr 11,000) was inactive but quickly regained activity upon removal of sodium dodecyl sulfate.

Enzyme Activation↗

PubMedDate not supplied

[Thermophillic sulfate-reducing bacteria from oil-bearing strata].

The paper describes pure cultures (strains 435 and 781) of thermophilic spore-forming sulphate-reducing bacteria isolated from oil strata. The strain 435 was classed as a new subspecies Desulfotomaculum nigrificnas subsp. salinus according to its morphological, physiological, and biochemical characteristics. The cells of the culture are rod-like, 2-5 mcm long and 0.9-1.3 mcm thick. Some cells are spindle-shaped. The cells have peritrichously arranged flagella. Spores are oval, located terminally or subterminally, slightly widening the cells. The culture is an obligate anaerobe. It grows in media containing sulphates and assimilates sodium salts of lactic, pyruvic, malic and formic acids, ethanol, and butanol. The cuture assimilates pyruvate in the absence of sulphates. Sulphate, sulphite and thiosulphate are electron acceptors when the culture grows on lactate. The organism can grow at 40-70 degrees C, the optimum temperature being 60 degrees C. It requires NaCl for growth, and can grow even in the presence of 4% NaCl in the medium. The optimum should contain 1% NaCl and microelements. Desulfoviridin is absent. The cells contain a cytochrome of the protoheme class. The content of G + C base pairs in DNA is 57.0 +/- 0.5 mol. %.

Anaerobiosis↗