Major Metabolic End Products Research Articles

Catabolism of aspartate and asparagine byBacteroides intermedius andBacteroides gingivalis

Aspartate as asparagine catabolism was studied in representative strains ofBacteroides intermedius strain T588 andB. gingivalis strain W83. Cell suspensions of both species deamidated asparagine. The enzyme asparaginase was constitutive and was unaffected by the addition of ammonium ions to the culture medium. The enzyme aspartase was not detected, but since malate dehydrogenase was known to occur and succinate was present as a major end product of metabolism, aspartate catabolism was postulated to occur via oxaloacetate, malate, and fumarate to succinate. All enzymes of this pathway were present in cell-free extracts, and some of the major properties of these enzymes were examined. The electron carriers cytochrome b and menaquinone-9 were present inB. gingivalis, whereasB. intermedius possessed cytochrome c and menaquinone-11. The membrane-bound enzyme fumarate reductase utilized NADH as an electron donor, but the reaction was inhibited by short wave ultraviolet radiation and 2-n-heptyl-4-hydroxyquinoline-N-oxide.

Sulfate: Turbidimetric and nephelometric methods

Publisher Summary This chapter discusses the turbidimetric and nephelometric methods for the estimation of sulfate. Inorganic sulfate is the major end product of sulfur metabolism in higher animals and is excreted mainly via the kidney. Its determination in urine thus allows the assessment of the dietary intake of sulfur amino acids in animals and human beings. Methods currently favored are based on the precipitation of sulfate with barium ions, but the old gravimetric procedures that are insensitive and tedious have been superseded by turbidimetric or nephelometric methods as described in the chapter. Turbidimetric or nepheiometric procedures require that certain precautions be taken. They should be performed under acid conditions to avoid interference from phosphate and preformed barium sulfate, and a stabilizing agent should be present in the assay to obtain adequate precision. The turbidimetric procedure is convenient for biological samples containing very little protein but a relatively high concentration of sulfate as in samples of urine. The nephelometric procedure, on the other hand, involves a deproteinization step and is applicable to samples of lower concentrations of sulfate, such as serum or plasma.

Cellodextrin utilization and beta-glucosidase production by Bacteroides polypragmatus.

Bacteroides polypragmatus, a mesophilic obligate anaerobe, was shown to simultaneously ferment glucose and cellobiose giving ethanol as a major metabolic end-product. A mixture of higher cellodextrins was also utilized. The bacterium produced a beta-glucosidase with a pI value of 4.2 and a molecular weight of approximately 100,000 daltons. The enzyme was intracellular and functioned optimally at pH 7. The Km values obtained with p-nitrophenyl-beta-D-glucoside and cellobiose as substrates were 0.73 mM and 100 mM, respectively. The enzyme was quite stable at elevated temperatures; in the presence of 10% glycerol (v/v), it had a half-life of 4 h at 55 degrees C. It was also stable during long-term storage at either 4 degrees C or -20 degrees C, provided that 10% (v/v) glycerol was added to preparations maintained at -20 degrees C.

Comparison of the distribution kinetics and metabolism to acid end-products of corticosterone and 11-deoxycorticosterone in BALB/c mice

The conversion of [4 14C]corticosterone([ 14C]B) and 11-deoxy-[1,2- 3H]corticosterone ([ 3H]DOC) to steroidal carboxylic acids was studied in the BALB/c mouse. There was rapid and preferential excretion of [ 3H]DOC metabolites into the gastrointestinal tract. Excretion of 14C through the kidney was higher than 3H excretion. Within minutes of intraperitoneal injection, levels of 3H and 14C in most organs reached their maximal levels and subsequently decreased in an exponential pattern. The majority of the organs took up 14C to a greater extent than 3H. Using tissue blood ratio of tracer (T/B) as criterion, it was found that liver, gall bladder, intestine, and kidney concentrated 3H and 14C-labeled steroid from blood. T/B for 3H exceeded that for 14C in the gastrointestinal tract. Abdominal fat preferentially took up [ 3H]DOC tracer, whereas [ 14C]B tracer was not taken up by this tissue. T/B was less than 1 for 3H and 14C in heart, thymus, spleen, brain, skeletal muscle and skin. In these organs uptake of B and its metabolites was greater than that of DOC and its metabolites. In liver, [ 14C]B and [ 3H]DOC were converted to carboxylic acid metabolites which accumulated in the intestine. The most abundant acid was 11β,20α-dihydroxy-3-oxo-pregn-4-en-21-oic acid from B. The acid metabolites of DOC were not identified. For both steroids, acids were major metabolic end-products.

High-performance liquid chromatographic determination of uric acid in soil

The large deposits of organic nitrogen in sea-bird rookeries may be responsible for an establishment of an ornithocoprophilous plant community. Uric acid, a major end-product of avian nitrogen metabolism, must play an important role in making this unique ecosystem. The acid was extracted from soil into potassium phosphate solution and separated from other components in the extract by high-performance liquid chromatography. Although the adsorption of the acid by soil prevents its quantitative recovery, the method is adequate to be applied to a study of nitrogen metabolism in rookeries. A large input of uric acid during the breeding season, its localization on the soil surface and its in situ decomposition were found to occur in the rookeries. The method is easily modified for preparative purposes, which makes it possible to perform isotopic analysis at a natural level and to elucidate further the nitrogen dynamics in the ecosystem.

LACTATE METABOLISM IN THE RUMEN OF SILAGE-FED SHEEP

Isotope kinetic studies in silage-fed sheep showed that while acetate was the major end-product of lactate metabolism, the molar proportion of propionate produced from lactate was higher than from other fermentable components of the silage. Lactate did not contribute directly to glucose synthesis but did via propionate. Key words: Sheep, lactate, silage

The anaerobic molluscan heart: Adaptation to environmental anoxia. Comparison with energy metabolism in vertebrate hearts

1.1. The hearts of many bivalve and gastropod molluscs are resistant to exposure to hypoxic and anoxic conditions.2.2. Glycogen and aspartate are simultaneously fermented leading to the accumulation of alanine, succinate and alanopine/strombine.3.3. Lactate is not a major end product of anaerobic metabolism in molluscan hearts.4.4. In contrast, vertebrate hearts respond to hypoxia by the fermentation of glycogen leading to lactate formation.5.5. There is some evidence for aspartate and glutamate breakdown in vertebrate hearts during anoxia. However, the quantitative contribution of this process to energy production is small.6.6. The differences in modes of energy production in molluscan and vertebrate hearts may reflect adaptations to long-term as opposed to short-term anoxia.

Microbial methanol formation: A major end product of pectin metabolism

Various pectinolytic strains ofClostridium, Erwinia, andPseudomonas species produced methanol as a major end product during growth on pectin but not on glucose or polygalacturonic acid. Pectin metabolism ofClostridium butyricum strain 4P1 correlated with a final product concentration of 16 mM at the end of growth, and a 1:1 stoichiometry for methanol production and percent initial substrate methoxylation. Growth on pectin was associated with high activity of pectin methylesterase and the absence of methanol consumption. The ecological significance of pectin metabolism and the establishment of microbial methylotrophic metabolism in nature is discussed.

Production of uric acid in the adult lung worm, Metastrongylus apri (nematoda: metastrongyloidea), from pigs

1.1. Uric acid was found to be a major end product of purine metabolism in adult Metastrongylus apri.2.2. The rates of production of radioactive uric acid during the incubation of intact worms with [8-14C]adenine, [8-14C]adenosine, [8-14C]hypoxanthine and [8-14C]guanine were measured.3.3. Most of the guanine, adenosine and hypoxanthine were catabolized directly but adenine was first converted to nucleotides before degradation.

Tryptophan metabolism during development of the stick insect,Carausius morosus Br. tissue distribution and interrelationships of metabolites of the kynurenine pathway

During larval development ofCarausius morosus kynurenic acid is the major end product of tryptophan metabolism. Tryptophan and kynurenic acid have been found in the fat body, haemolymph and gut contents but only traces of kynurenine have been detected. The ommochromes ommin and xanthommatin are formed in relatively small amounts in the epidermis during larval development. 3-hydroxykynurenine was found only in the epidermis, the site of ommochrome deposition.

The quantitative role of the kidneys in the in vivo metabolism of mevalonate.

The roles of the sterol and nonsterol pathways in the metabolism of circulating mevalonate have been estimated in the intact rat. On an average, the sterol pathway accounts for 74 per cent of the mevalonate metabolized, while the nonsterol, or shunt, pathway is responsible for 26 per cent of the mevalonate metabolized in the whole animal. The contribution of the kidneys to each of these processes was evaluated by two approaches. First, the localization of labeled sterols and sterol precursors derived from [14C]mevalonate was determined in each of the major tissues of the body and, second, the effect of nephrectomy upon mevalonate metabolism by the sterol and shunt mechanisms was examined. The results confirm our earlier conclusion that the kidneys represent the primary tissue site of conversion of circulating mevalonate to sterols and sterol precursors. In the present study, it was shown that by 6 h after administration of [14C]mevalonate, the major end product of mevalonate metabolism in the kidneys is cholesterol and that, moreover, the kidneys are responsible for most of the cholestreol synthesized in the intact animal from injected mevalonate. Following nephrectomy, the extrarenal tissues can readily assume the dominant role normally played by the kidneys in synthesizing cholesterol and other sterols from circulating mevalonate. The major observation of the present study is that the kidneys represent the primary site of mevalonate metabolism by the shunt pathway, in that nephrectomy results in approximately a 60 per cent decrease in the mevalonate metabolized by the shunt pathway. These studies, therefore, reinforce and expand the evidence that the kidneys represent the most important single tissue site for the metabolism of circulating mevalonate.

Comparative Anti-Inflammatory, Analgesic, and Antipyretic Activities of 7-Chloro-3,3a-dihydro-2-methyl-2H,9H-isoxazolo-(3,2-b)(1,3)-benzoxazin-9-one and 5-Chlorosalicylic Acid in Rats

Evidence is presented which indicates that 7-chloro-3,3a-dihydro-2-methyl-2H,9H-isoxazolo-(3,2-b)(1,3)-benzoxazin-9-one (I) and 5-chlorosalicylic acid, its major metabolic end-product, are equally effective as anti-inflammatory and antipyretic agents, while the former is a somewhat more effective analgesic than its metabolite in the rat. However, at the equimolar doses used in this study, I is not ulcerogenic, while 5-chlorosalicylic acid does possess this untoward effect in the fasted rat. Moreover, the LD50 for 5-chlorosalicylic acid (261.0 mg/kg) is approximately 6.5 times less than that of I (1710.0 mg/kg) in the nonfasted rat. These results support the postulation that 5-chlorosalicylic acid is most likely responsible for the pharmacological activity displayed by I; i.e., the latter acts as a carrier or delivery system, allowing attenuation of the toxic properties of its active metabolite.

Comparative metabolism of 3-amino-1,2,4-triazole

1. The metabolism of the herbicide amitrole (3-amino-1,2,4-triazole) in rats has been studied. Unaltered amitrole and three metabolites are present in the urine of treated animals. 2. The amitrole metabolites have been compared with those formed by beans and Escherichia coli. The two major metabolic end-products of amitrole have been found to be similar in these organisms, and a third one is common to both beans and rats. 3. One of the major metabolites has been isolated and recrystallized and some structural data have been obtained. Its infrared spectrum as well as its acid-base behaviour are compatible with the structure of amitrolylalanine proposed by Massini (1963). The results are also discussed in terms of the possible metabolic pathways leading to amitrolylalanine formation.

Altered Metabolism in a Streptococcus lactis C2 Mutant Deficient in Lactic Dehydrogenase

A spontaneous mutant of Streptococcus lactis C2 which formed abnormally large colonies on agar medium was isolated. The mutant grew as rapidly in broth or milk as the parent culture but was slower in acid production. When grown in complex broth at 32C, the parent culture lowered the pH to 4.8 in 8h whereas the mutant required 15h to produce pH 5.0. Mutant cells consumed 6 times as much oxygen as S. lactis C2 and produced carbon dioxide derived primarily from carbons 3 and 4 of glucose. The mutant was defective in lactic dehydrogenase possessing only .1 units as compared to 138 units for S. lactis C2. This enzymatic defect caused the mutant to produce about 1,000μg/ml of Westerfeld positive material when grown in broth. S. lactis C2 produced only trace amounts. Although acetoin was a major metabolic end-product in the mutant, diacetyl was also detected. The results indicate a type of mutation which could be responsible for the spontaneous occurrence of aroma and carbon dioxide producing strains of S. lactis in dairy starter cultures.

The metabolism of carbohydrate by pleomorphic African trypanosomes

1.1. Monomorphic Trypanosoma brucei and T. rhodesiense metabolize glucose via the glycolytic pathway, producing pyruvate as the major metabolic end-product with a small amount of glycerol and no carbon dioxide, acetate or succinate.2.2. Under in vitro conditions, pleomorphic strains of T. rhodesiense produce significant quantities of succinate, acetate and carbon dioxide.3.3. The time course of metabolite production has been used to evaluate the significance of acetate, succinate and carbon dioxide formation in vivo and indicates that succinate production is induced by in vitro incubation.4.4. Oxidative decarboxylases for pyruvate and α-oxoglutarate are confined to the short stumpy forms of pleomorphic infections. However, the tricarboxylic acid cycle has minimal activity in these organisms due to limiting levels of citrate synthase (E.C. 4.1.3.7) and succinate dehydrogenase (E.C. 1.3.99.1).5.5. l-Glycerol-3-phosphate oxidase is the principal terminal oxidase of both long slender and short stumpy forms of T. rhodesiense.

Metabolic sensitivity of Fasciola hepatica to mammalian hormones in vitro

1. 1. Intact liver fluke ( Fasciola hepatica) and liver fluke homogenates were incubated with mammalian hormones in the presence of a range of radioactively labelled substrates. The incorporation of radioactivity into some of the major metabolic end products (carbon dioxide, lipid and glycogen) was used to assess the sensitivity of the parasite to the hormones. 2. 2. No significant effects were observed with thyroxine, histamine, epinephrine, norepinephrine, progesterone, testosterone and hydrocortisone. A response was detected in the presence of 5-hydroxytryptamine. 3. 3. The difficulty of interpretation of the negative result obtained is discussed.

The relationship of some intermediary metabolites to the production of volatile fatty acids by adult Fasciola hepatica

1.1. The degradation of pyruvate to volatile fatty acids by Fasciola hepatica in vitro was determined, and the incorporation of labelled pyruvate into acetic and propionic acids was demonstrated.2.2. The major end product of 14C-pyruvate metabolism was found to be acetic acid with minor amounts of radioactivity in propionic acid.3.3. The incorporation of 14CO2 into propionate was demonstrated in vitro. However no incorporation into acetate could be shown.4.4. Exogenous succinate or citrate were found to be minor contributors to propionate and acetate production.5.5. Volatile fatty acid production was found to be stimulated by serotonin in the presence of glucose or pyruvate, and in the absence of exogenous substrates.

Biochemistry of the dog hookworm: II. Nature and origin of the excreted fatty acids

The fermentation of d-glucose- 14C by adult Ancylostoma caninum leads to the production of radioactive acetate, propionate, and carbon dioxide. The propionate appears to be a major end product of glucose metabolism. A large proportion of the glucose is metabolized to one or more nonvolatile compounds. The branched chain acids, isobutyric and isovaleric, are also excreted by A. caninum. Isovaleric acid was identified by infrared spectroscopy. The branched chain acids are excreted in a fixed proportion to the total volatile acids produced. The branched chain acids are not derived from glucose but can be readily produced from amino acids. l-Valine is the precursor of isobutyric acid, and l-leucine is the precursor of isovaleric acid. The labeling of propionate and carbon dioxide, but not acetate, when l-valine- 14C and l-leucine- 14C were metabolized indicates that propionate formation may occur via a carbon dioxide fixation pathway. A “Pasteur effect” was observed in fermentation experiments with A. caninum.

Relationship of n-Valeric Acid Content and Intensity of Infection in Trichinous Hogs

Muscles from 12 moderately or heavily infected hogs contained relatively large amounts (0.25 to 13.70 Atg/g) of n-valerie acid, an acid rarely found in vertebrate tissues. The n-valeric acid content in general, correlated well with the larval counts in the diaphragm, tongue, and psoas musclescorrelation coefficients of 0.81, 0.84, and 0.82, respectively, were obtained. Only trace amounts of the acid were found in the visceral organs of the infected hogs. Trace amounts were also found in the muscles and organs of hogs with infections of less than 21 days duration, in hogs with no known helminth infections, and in hogs with ascaridand kidney worm-infections. The level of n-valeric acid in infected muscle per larva counted was about twice that per decapsulated larva or per isolated cyst from the same muscle. The production of the volatile fatty acid, n-valeric, in animal metabolism is evidently uncommon (Bueding and Most, 1953; von Brand et al., 1952). Bacteria and molds produce this acid (bacteria, Barker and Haas, 1944; Bornstein and Barker, 1948; Dehority et al., 1958; Nakae and Elliot, 1965; Yoshiaki and Karashimada, 1966; Serzedello, 1967; molds, Leal et al., 1966; Nishibe et al., 1966). Consequently, it occurs in swine feces where the diet controls its proportion among the volatile fatty acids (Friend et al., 1962). In the parasitic helminths, this acid was probably first observed by Bunge (1889) when he was studying anaerobic fermentation in Ascaris lumbricoides and A. megalocephala (= Parascaris equorum). Bunge found that the products of the fermentation process were CO2 and a strong-smelling, volatile fatty acid that he did not identify. Weinland (1901, 1902, 1903), with objectives similar to those of Bunge and using the same nematodes, identified the strong-smelling component as valeric acid and as a major end product of carbohydrate metabolism. Later, other isomers of valeric acid and other fatty acid components were identified in A. lumbricoides (Bueding and Yale, 1951; Bueding, 1953). In in vitro studies concerning carbohydrate metabolism of the larvae of Trichinella spiralis, von Brand et al. (1952) found n-valeric acid to be a major end product. The present work was undertaken to determine if n-valerie acid could be detected in Received for publication 10 October 1969. certain muscles and organs of trichinous hogs to ascertain if such a determination could be used to diagnose trichinosis in swine. MATERIALS AND METHODS The larvae and cysts of Trichinella spiralis utilized in this study were obtained from hogs infected with a strain of the parasite that had been maintained in swine at this laboratory for more than 30 years. In all experiments, each hog to be infected was fed trichinous pork at the rate of 500 larvae per pound of live body weight. The experimental animals were fed a well-balanced mash feed and, in some cases, shelled corn. Experimental animals All experimental hogs were raised worm-free until weaned, in concrete pens that were washed daily, as recommended by Spindler (1942). The 11 noninfected controls as well as 10 of the 15 infected hogs were maintained in such pens until slaughter; 5 of the infected hogs were maintained in a fenced hog lot. Spot checks at slaughter for trichurids and asearids in the controls and experimentally infected trichinous hogs were negative. The duration of the infections in the 15 infected hogs ranged either from 45 days to 8 months or from 11 to 20 days. Also, 3 ascaridand 2 kidney worm-infected pigs, which were used in other experiments, were used as helminth-infected, nontrichinous controls. At necropsy, the hogs weighed from 95 to 400 lb. Preparation of tissue samples The following tissue samples were taken from each of the 15 infected and 11 control hogs at slaughter: diaphragm (pillars), tongue, psoas, intercostals, loin, heart, duodenum, liver, and kidney. The samples were washed and ground separately in an electric food chopper. The ground samples of each tissue were well mixed and 2 or 3 10-g portions removed within 3 hr after the death of the donor, and processed for examination

Carbohydrate metabolism in Babesia rodhaini: Differences in the metabolism of normal and infected rat erythrocytes

Glucose metabolism in rat erythrocytes appeared to be qualitatively similar to that reported for the erythrocytes of other mammalian species, although the rates of oxygen uptake and glucose utilization were greater than those of the erythrocytes of larger animals. Erythrocytes incubated with excess glucose as substrate had a high equilibrium ratio of pyruvate to lactate, which may have been due to high rates of NADH-dependent methemoglobin reductase activity. When rat erythrocytes were deprived of substrate they continued to respire at a constant rate for at least 2 hours; there was a marked accumulation of pyruvate during this period. Increased reticulocyte percentage in rats' blood led to increased respiratory and glycolytic rates. There was a high positive correlation between reticulocyte percentage and oxygen uptake. B. rodhaini-infected rat erythrocytes exhibited rates of oxygen uptake and glycolysis far in excess of normal rat erythrocytes. However, the ratio of oxygen uptake to glucose utilization was no greater than that in normal rat erythrocytes, and less than that reported for erythrocytes infected with Plasmodium spp., suggesting, therefore, a greater dependence of B. rodhaini on anaerobic metabolism. Lactate was always the major end product of glucose metabolism in B. rodhaini-infected red blood cells. The glycolytic ratios, and the percentages of glucose which could be accounted for as oxygen uptake, lactate, and pyruvate, indicated a greater use of glucose for synthetic purposes in infected cells when compared with normal rat erythrocytes. When the parasite was within its host erythrocyte, it continued to respire and produce lactate and pyruvate for some hours in the absence of added substrate.