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PubMed2005

Migrastatin and dorrigocins are shunt metabolites of iso-migrastatin.

Fermentation of Streptomyces platensis NRRL18993 typically accumulated migrastation (1), dorrigocin A (2) and B (3), and 13-epi-dorrigocin A (5). Supplement of XAD-16 resin to the fermentation, in contrast, resulted in exclusive production of iso-migrastatin (4). In vitro studies showed that 1, 2, 3, and 5 are stable in aqueous solution but 4 undergoes rapid conversion into 1, 2, 3, and 5 under the same condition. These results revealed that 4 is the only bona fide natural product biosynthesized by S. platensis, and 1, 2, 3, and 5 are shunt metabolites of 4. This study also established the stereochemistry of 2-5 with the exception of C-11 for 3 and 4. A mechanism for H2O-mediated regio- and stereospecific rearrangement of 4 to 1, 2, 3, and 5 is proposed and supported by incorporation of 18O from H218O.

Chromatography, High Pressure Liquid↗
PubMed2003

Raw potato starch and short-chain fructo-oligosaccharides affect the composition and metabolic activity of rat intestinal microbiota differently depending on the caecocolonic segment involved.

AIMS: In vitro studies have suggested that fructo-oligosaccharides (FOS) and resistant starch (two fermentable non-digestible carbohydrates) display different fermentation kinetics. This study investigated whether these substrates affect the metabolic activity and bacterial composition of the intestinal microflora differently depending on the caecocolonic segment involved. METHODS AND RESULTS: Eighteen rats were fed a low-fibre diet (Basal) or the same diet containing raw potato starch (RPS) (9%) or short-chain FOS (9%) for 14 days. Changes in wet-content weights, bacterial populations and metabolites were investigated in the caecum, proximal and distal colon and faeces. Both substrates exerted a prebiotic effect compared with the Basal diet. However, FOS increased lactic acid-producing bacteria (LAPB) throughout the caecocolon and in faeces, whereas the effect of RPS was limited to the caecum and proximal colon. As compared with RPS, FOS doubled the pool of caecal fermentation products, while the situation was just the opposite distally. This difference was mainly because of the anatomical distribution of lactate, which accumulated in the caecum with FOS and in the distal colon with RPS. Faeces reflected these impacts only partly, showing the prebiotic effect of FOS and the metabolite increase induced by RPS. CONCLUSIONS: This study demonstrates that FOS and RPS exert complementary caecocolonic effects. SIGNIFICANCE AND IMPACT OF THE STUDY: The RPS and FOS combined ingestion could be beneficial by providing health-promoting effects throughout the caecocolon.

Animals↗
PubMed1995

Fly ash, basic slag and Glenbrook slag toxicity in cattle.

In case studies of two herds of dairy cattle, a herd of 270 Friesian cows and heifers and a herd of 256 Jersey cows and heifers experienced high death rates and a protracted course of illness when fly ash was broadcast as a fertiliser over the farms at a rate of about 1 tonne per hectare. The onset of weakness, lassitude and diarrhoea in the cattle first appeared within 4-5 days and both fertility and milk fat production fell off dramatically. In the following 7 months, 51 animals died in the Friesian herd and a similar death rate and performance losses occurred in the Jersey herd. Emaciation and hepatic fibrosis were consistently reported from autopsies. Heavy metal poisoning was eliminated from the possible diagnosis. The slags and fly ash used as fertiliser had several features in common. They contained 29-40% calcium as calcium oxide, 15-35 grams of iron per kg, they were all extremely alkaline and concentrations of lead and arsenic were low. The effect of adding fly ash to rumen contents in vitro was to raise the pH to greater than 7.2, at which point fermentation was markedly-reduced.

Journal Article↗
PubMed2000

A cyclooxygenase-2 (COX-2) selective non-steroidal anti-inflammatory drug enhances the growth inhibitory effect of butyrate in colorectal carcinoma cells expressing COX-2 protein: regulation of COX-2 by butyrate.

Epidemiological, clinical, animal and laboratory studies have all provided evidence for the protective effects of non-steroidal anti-inflammatory drugs (NSAIDs), such as aspirin, against colorectal cancer. The main established target for NSAID action is cyclooxygenase (COX) and the inducible isoform, COX-2, is up-regulated in colorectal cancer. Rat intestinal epithelial cells transfected with a COX-2 expression vector have previously been found to be resistant to butyrate-induced apoptosis. Butyrate, a by-product of dietary fibre fermentation, is known to induce differentiation and apoptosis in colorectal tumour cells in vitro. In recent years there has been considerable interest in the possible role of dietary fibre/resistant starch in the prevention of colorectal cancer. In this study we investigated whether inhibition of COX-2 with a highly selective COX-2 inhibitor (NS-398) would sensitize human colorectal carcinoma cells to the growth inhibitory effect of butyrate. HT29 and S/KS colorectal carcinoma cell lines were treated for 72 h with 2 mM butyrate and/or 10 microM NS-398. Addition of 10 microM NS-398 alone (to inhibit COX-2 activity) did not result in detectable growth inhibition in either of the cell lines. NS-398 enhanced sensitivity to the growth inhibitory effect of butyrate in HT29 cells expressing COX-2 protein. In contrast, NS-398 did not sensitize S/KS cells lacking detectable COX-2 protein and function (as determined by prostaglandin E(2) production) to the growth inhibitory effect of butyrate. In addition, we report that butyrate treatment of carcinoma (HT29) and adenoma (PC/AA/C1) cells leads to up-regulation of COX-2 protein. Thus NS-398 only appears to sensitize human colorectal carcinoma cells expressing COX-2 protein to the growth inhibitory effect of butyrate. As COX-2 is up-regulated in colorectal carcinogenesis, this could have important implications for the selective inhibition of cells expressing COX-2 protein over those lacking COX-2 protein expression and for dietary modification to be considered alongside NSAIDs in the prevention, and possibly treatment, of colorectal cancer.

Animals↗
PubMed1998

The chondramides: cytostatic agents from myxobacteria acting on the actin cytoskeleton.

BACKGROUND: Chondramides are cyclodepsipeptides produced by strains of the myxobacterium, Chondromyces crocatus. These peptides, which have been reported to inhibit yeast and mammalian cell proliferation, are related to jasplakinolide, which has been isolated from marine sponges of the genus Jaspis and has been shown to interfere with the actin cytoskeleton (a structural component of cells that helps maintain their shape and is involved in processes, such as cell division and cell locomotion). We studied the effects of the chondramides (A, B, C, and D) on tumor cell growth, on cytoskeletal structure, and on actin polymerization in vitro and compared these effects with those of cytochalasin D and jasplakinolide. METHODS: Cell proliferation was measured by means of tetrazolium salt reduction assays. Effects on the cytoskeleton were studied by use of fluorescence techniques, and actin polymerization in vitro was measured by means of viscosimetry. RESULTS: Proliferation of tested tumor cell lines was inhibited by the chondramides. Concentrations that inhibited proliferation by 50% (IC50 values) ranged from 3 to 85 nM and were of the same order of magnitude as those found for cytochalasin D and jasplakinolide. Fluorescence staining of potoroo cells incubated with chondramides A and B showed that organization of the actin cytoskeleton was disrupted; however, the microtubule system was not affected. Viscosimetric measurement showed that, depending on the experimental conditions, chondramide A induced or accelerated actin polymerization in vitro. CONCLUSION: The chondramides--unlike jasplakinolide--can be produced in large amounts by fermentation, and, similar to jasplakinolide, they appear to have antiproliferative activity against carcinoma cell lines by targeting the actin cytoskeleton.

Actins↗
PubMed1997

Inhibition of anchorage-independent growth of tumor cells by IT-62-B, a new anthracycline.

IT-62-B, a new anthracycline isolated from fermentation broths of Streptomyces sp. IT-62, reversed certain tumor cell phenotypes in vitro including some of human origin. The observed normal phenotypes were anchorage dependence of cell growth, flattened cell morphology and restoration of actin stress fibers. The extent of the anchorage dependence of cell growth induced by IT-62-B was generally greater than that by doxorubicin or pirarubicin. The cell-flattening effect of IT-62-B on cells of T24 (human bladder), but not on C-33A (human cervix), accompanied inhibition of fos gene expression. T24 cells, once flattened by IT-62-B, retained their flat morphology even in drug-free, fresh medium and eventually died in several days. IT-62-B, unlike doxorubicin, only slightly inhibited the topoisomerase II reaction in vitro and DNA synthesis in isolated cell nuclei.

Actins↗
PubMed1998

Recent developments in tetracycline antibiotics.

The rapid emergence of pathogenic bacteria resistant to tetracyclines and other currently available antibiotics has caused serious concern among medical professionals. It has heightened resurgent interest in studying the mechanisms of resistance and in developing new antibiotics. A comprehensive review has outlined the developments of tetracyclines prior to 1980 [47]. This review will highlight the pertinent advances in the tetracycline field during the last two decades, including recent progress on elucidating the mechanisms of resistance, and the development of novel tetracyclines to combat bacterial resistance. Most of the new tetracycline derivatives described in this review have been either prepared semisynthetically or isolated from fermentation. In the semisynthetic area, efflux inhibitors that are effective in an in vitro model have been identified. A new class of tetracyclines, named glycylcyclines has been the subject of numerous reports, and will be the major focus of this review. The glycylcyclines are currently the only derivatives that exhibit antibacterial activity comparable to that of the early tetracyclines when they were first introduced. These compounds show potent activity against a broad spectrum of Gram-positive and Gram-negative bacteria, including strains that carry the two major tetracycline-resistance determinants, efflux and ribosomal protection. Two of the glycylcycline derivatives. DMG-MINO and DMG-DMDOT, have been studied by several groups of investigators against a large number of clinical pathogens isolated from various sources. The spectrum of activity of these compounds includes organisms with resistance to antibiotics other than tetracyclines, e.g., methicillin-resistant staphylococci, penicillin-resistant streptococcus pneumoniae, and vancomycin-resistant enterococci. Their in vitro, as well as in vivo activity against bacteria with characterized tetracycline- or minocycline-resistant elements will be summarized. The structure-activity relationships of glycylcyclines and their mode of action will also be discussed.

Anti-Bacterial Agents↗
PubMed1989

[Non-cariogenicity of maltitol in vitro and animal experiments].

The caries inducing activity of maltitol was examined in in vitro and animal experiments. Fifteen strains of oral streptococci were not able to ferment maltitol. Also, maltitol inhibited the glucan synthesis from sucrose by GTases from mutans streptococci. However, the inhibition of growing-cell adherence was not so marked with the addition of maltitol to sucrose culture. In the animal experiments, maltitol did not induced dental caries in rats infected with mutans streptococci. Furthermore, maltitol significantly inhibited the sucrose-induced dental caries in rats infected with S. sobrinus 6715. However, there was no significant difference in rats infected with S. mutans MT8148R. These results indicated that maltitol is useful as a non-cariogenic sucrose substitute.

Animals↗
PubMed1988

Antitumor activity and hematotoxicity of a new, substituted dihydrobenzoxazine, FK973, in mice.

FK973, a new, substituted dihydrobenzoxazine (11-acetyl-8-carbamoyloxymethyl-4-formyl-14-oxa-1,11- diazatetracyclo[7.4.1.0.0]tetra-deca-2,4,6-trien-6,9-diyl diacetate), was obtained by chemical modification of a novel antibiotic which was isolated from the fermentation products of Streptomyces sandaensis No. 6897. FK973 had cytotoxic effects against in vitro cultured human and murine tumor cells. FK973 in doses of 0.032-5.6 mg/kg (i.p.) had stronger antitumor activities and higher chemotherapeutic ratio than mitomycin C against such murine ascitic tumors as P388 and L1210 leukemia, B16 melanoma, M5076 reticulum cell sarcoma of ovarian origin, Colon 26 carcinoma, Ehrlich carcinoma, and MH134 hepatoma. In tests against murine and human solid tumors implanted s.c. in normal mice and nude mice, respectively, FK973 (i.v.) inhibited growth of murine tumors (M5076 sarcoma, Colon 38 carcinoma, B16 melanoma, and Lewis lung carcinoma) by 66-100% and human tumors (LX-1 lung, MX-1 mammary, and SC-6 stomach carcinoma) by 84-99%. In studies with drug-resistant P388 leukemia, FK973 was also effective against vincristine-resistant P388, moderately effective against mitomycin C (MMC)- and adriamycin-resistant P388, and partially effective against cyclophosphamide-resistant P388 cells in mice. Leukopenic effects of FK973 and MMC in mice were comparable at doses which gave antitumor activity almost equally. FK973 had no effect on the numbers of platelets and red blood cells, whereas MMC markedly decreased both. FK973 decreased the numbers of colony forming units in spleen and in culture and the effect was less than that of MMC. Therefore, FK973 may give weaker myelosuppression than MMC. The results suggest that FK973 will be a beneficial drug for the treatment of cancer.

Animals↗
PubMedDate not supplied

In vitro and in vivo characterization of avian Escherichia coli. I. Serotypes, metabolic activity, and antibiotic sensitivity.

Over a 2 1/2-year period (January 1981 to June 1983), 177 Escherichia coli isolates were obtained from 145 field-reared broiler flocks in the Delmarva peninsula, and 20 were obtained from clinically normal day-old hatchery chickens representing an additional 17 flocks in Delmarva. Ninety-one isolates obtained from the field-reared birds between 2 and 8 weeks of age were associated with complicated air-sac disease. Serotyping efforts demonstrated a predominance of O2, O35, and O78 serogroups and a large number of untypable isolates. More than 50% of the isolates in each of the three dominant serogroups were collected from broilers with colibacillosis, but they were never detected in the yolk-sac samples of clinically normal day-old hatchery chickens. In vitro biochemical characterization of the E. coli isolates revealed variable rates of carbohydrate fermentation and amino acid decarboxylation. No common characteristics appeared to be shared by the predominant serogroups isolated from clinically affected birds, although several serogroup-specific reactions were noted. The majority of the isolates were sensitive to chloramphenicol, gentamicin, nalidixic acid, ormethoprim-sulfadimethoxine, spectinomycin, neomycin, and ampicillin. About half of the isolates were sensitive to nitrofurantoin and the sulfa compounds. Less than 25% of the isolates were sensitive to streptomycin, erythromycin, tetracyclines, novobiocin, penicillin, bacitracin, and lincomycin.

Animals↗
PubMed1995

The application of a semi-continuous colon simulation technique (Cositec) for studying the effects of clindamycin on microbial hindgut metabolism.

The semi-continuous colon simulation technique was used as an experimental in vitro model to study potential effects of clindamycin application on basic parameters of microbial hindgut metabolism. Hindgut contents from fistulated pigs kept on conventional diets were used as substrates for incubation. Measurements on reproducibility demonstrated the colon simulation technique as a suitable in vitro method to characterize microbial hindgut metabolism and those factors potentially influencing microbial fermentation. Application of clindamycin resulted in disturbances of microbial steady state metabolism and time- and dose-dependent decreases of SCFA production rates with significant reductions of molar butyrate proportions to almost zero. These effects can be related to the onset of functional disturbances of hindgut function under in vivo conditions since butyrate is an essential trophic factor for colonocytes. In addition, clindamycin induced increases in lactate production with a shift towards D-lactate. The relevance of these changes for hindgut function have to be elucidated in further experiments.

Animals↗
PubMed1995

Laxatives and intestinal epithelial cells: a morphological study of epithelial cell damage and proliferation.

Experimental studies indicate that anthranoid laxatives may induce epithelial damage. In addition they induce the release of prostaglandins. Epithelial cell damage and release of prostaglandins are two pathways by which epithelial cell proliferation could be influenced. Other laxatives including fermentable laxatives like lactulose may also influence large intestine cell proliferation by the trophic effect of the fermentation products such as short chain fatty acids. For these reasons an in vitro study was performed on human intestinal epithelial cells in culture to investigate the direct damaging effect of rhein anthrone. Ultrastructural examination showed a dose dependent direct damage. In addition an in vivo study in rats was performed to compare the short and long term effect of sennosides, bisacodyl, sodium picosulphate and lactulose on epithelial cell proliferation in the ileum and large intestine. Cell proliferation was examined by the BrdUrd labeling technique after 2, 6 and 12 weeks of continuous treatment. Studies in control animals show that the Labeling Index (LI) is higher in the caecum compared with other segments of the colon, and higher in the ileum than in the colon. Treatment with sennosides, bisacodyl and sodium picosulphate does not influence the LI in the ileum and induces no statistically significant increase of the LI when the treated groups are compared with the control group at the end of the study. The proliferative pattern along the crypts remains unchanged with all the laxatives throughout the study. It appears therefore that 'contact' laxatives have no major influence on ileal and colonic epithelial cell proliferation.

Animals↗
PubMed1976

Application of synthetic estrogen in sheep.

Thirty castrated crossbreed lambs of 4 months age were divided into three groups. DES pellets (6 mg) were implanted subcutaneously in lambs of groups II and III, respectively at 4 and 7 months of age whereas those in group I served as controls. The lambs were fed on a dry fattening ration during a period of 29 weeks after which two lambs of each group were slaughtered and three lambs were also used in nitrogen balance studies. The body gains of lambs implanted with DES at 4 months of age were the highest. The growth promoting effect of the hormone in these lambs was significant during a period of 13 weeks after the implantation. The dietary nitrogen retained by treated lamb was significantly higher. The dressing percentage and weights of wholesale cuts in lambs implanted with DES were similar to those of control lambs. However, the percentage of meat in the lambs treated at 4 months of age was the highest. The protein and moisture contents of the tenth rib of these lambs were greater and the fat contents were lower than in the control animals. No DES residual activity was ever noted in the livers of slaughtered lambs. Effect of DES Implantation on Body Components. Six 2 year old Egyptian rams were used in a 2-month experiment, the duration being divided into three successive intervals. The 1st period served as a control. At the beginning of the 2nd period, DES was implanted subcutaneously. Total body water was measured using tritiated water, total muscle mass was determined by the creatinine excretion during 24 hrs, lean body mass, body rat, and nitrogen balance, were measured during the last 5 days of each experimental period. DES implantation increased the body weight of the ram by 10.4% and caused no significant change in total body water, body ash, or total muscle mass. However, body fat increased significantly. The efficiency of nitrogen utilization also increased significantly although nitrogen intake did not change. The maximum effects of DES were observed at the end of the second experimental period. Effect of Some Estrogens on Rumen Metabolism. Three DES treated and three untreated cross bred Egyptian rams were used for studying the effect of DES on rumen microorganisms. Ruminal activity, judged by the diurnal concentrations of volatile fatty acids (VFA) and ammonia-N determined 3 and 6 weeks after DES implantation, was greater in treated animals. When rumen contents from fistulated sheep were incubated in vitro with stilbestrol dipropionate (SDP), DES, and Estradiol dipropionate (EDP), a significant increase in the number and size of rumen ciliate protozoa was observed. The extent of increase varied according to the type and concentrations of added hormones and type of rumen protozoa. Further in vitro experiments indicated that the addition of DES, SDP, and EDP promoted the fermentation of starch by washed suspensions of mixed populations of ciliate protozoa. EDP seemed to show the greatest effect in stimulating VFA production by the protozoal cells.

Anabolic Agents↗
PubMed1995

In vitro ouabain-sensitive respiration and protein synthesis in rumen epithelial papillae of Hereford steers fed either timothy hay or timothy hay supplemented with cracked corn once daily.

Rumen epithelial papillae samples, acquired from the Central sac of six adult Hereford steers (585 +/- 17 kg) fed either timothy hay plus soybean meal (T) or timothy hay plus cracked corn and soybean meal (TC) once daily (0900), were used in a crossover design to study the daily pattern of O2 consumption and protein synthesis. Tissue samples were acquired at 0900, 1200, 1800, and 2400 over a 10-d sampling period. Additionally, ruminal fermentation characteristics (pH, ammonia, VFA, and osmolality) were measured at 0430, 0900, 1030, 1200, 1500, 1800, 2100 and 2400. Total, ouabain-insensitive (OIO2), and cycloheximide-insensitive (CIO2) O2 consumption were greatest at 2400 (P < .01, P < .06, and P < .02 respectively). Additionally, steer fed TC had a greater CIO2 at 2400. In conjunction with a temporal effect on ruminal fermentation patterns, after an initial decline, there was an increase (P < .05) in total O2, ouabain-sensitive (OSO2), OIO2, and CIO2 consumption throughout the day. Fractional rates of protein synthesis were not different at any time point between diets. Rumen epithelial metabolism exhibits a temporal pattern in cattle fed once daily.

Ammonia↗
PubMed1986

Influence of the oropharyngeal microflora on the measurement of exhaled breath hydrogen.

We investigated the possible contribution made by oropharyngeal microfloral fermentation of ingested carbohydrate to the generation of the early, transient exhaled breath hydrogen rise seen after carbohydrate ingestion. Ten subjects ate or were sham fed carbohydrate-containing meals with and without prior chlorhexidine mouthwash during serial collection of exhaled breath and mouth hydrogen samples. Meal ingestion and sham feeding both induced significant (p less than 0.01) elevations of breath and mouth hydrogen that were virtually abolished by prior chlorhexidine mouthwash. In 7 subjects, delivery of the meal directly into the stomach via an orogastric tube did not cause a breath or mouth hydrogen rise. Oral contents incubated anaerobically in vitro with carbohydrate generated hydrogen that was again inhibited by chlorhexidine. These studies indicate that fermentation of ingested carbohydrate by oropharyngeal bacteria can contribute significantly to measured breath hydrogen values soon after meal ingestion, and may introduce avoidable error into the interpretation of serial breath hydrogen data.

Adult↗
PubMed1997

Effects of ruminally degradable nitrogen intake and in vitro addition of ammonia and propionate on the metabolic fate of L-[1-14C]alanine and L-[15N]alanine in isolated sheep hepatocytes.

Isolated hepatocytes prepared from sheep fed a basal diet (bromegrass hay-corn, 50:50 wt/ wt, as-fed basis) with or without urea were used to determine the effects of added ammonia (as NH4Cl) and propionate on the partitioning of C from 1.25 mM L-[1-14C]alanine between oxidation and gluconeogenesis, and the flux of 15N from 1.25 mM L-[15N]alanine to [14N15N]urea and [15N15N]urea. Hepatocyte suspensions were incubated with NH4Cl (0, .31, .63, and 1.25 mM) and (or) propionate (0, .31, .63, and 1.25 mM) in the presence of either 1.25 mM L-[15N]alanine or 1.25 mM L-alanine plus 18.5 kBq of L-[1-14C]alanine. Feeding dietary urea did not affect [1-14C]alanine oxidation to 14CO2 (P = .601), or its conversion to [14C]glucose (P = .576) by isolated hepatocytes. Increasing in vitro concentrations of NH4Cl and propionate between 0 and 1.25 mM reduced [1-14C]alanine oxidation to 14CO2 (P < .001). Increasing NH4Cl concentration between 0 and 1.25 mM reduced [1-14C]alanine conversion to [14C]glucose in isolated hepatocytes (P < .001), whereas addition of propionate between 0 and 1.25 mM stimulated production of [14C]glucose from [1-14C]alanine (P < .001). Feeding urea did not affect in vitro rates of total urea production (P = .655) but increased the production of [14N15N]urea and [15N15N]urea (P < .05). Addition of NH4Cl increased total urea, [14N15N]urea, and [15N15N]urea production (P < .001), but reduced 15N isotopic enrichments of [14N15N]urea and [15N15N]urea (P < .001). Increasing propionate concentration between 0 and 1.25 mM reduced total urea production (P < .001), but [14N15N]urea and [15N15N]urea production was reduced only at 1.25 mM propionate (P < .001). We conclude that NH3 detoxification by isolated sheep hepatocytes increases amino acid deamination and this might have implications for nitrogen retention in ruminants consuming diets that promote considerable NH3 absorption from the digestive tract.

Alanine↗
PubMed2005

Influence of living and autoclaved yeasts of Saccharomyces boulardii on in vitro ruminal microbial metabolism.

Experimental data on the effects of Saccharomyces boulardii on rumen microbial metabolism are scarce. The aim of this study was to examine whether S. boulardii had an effect on parameters of rumen microbial metabolism at different dosages and whether the yeast would be suitable as a probiotic agent for ruminants. To test whether the potential positive effects of S. boulardii could be attributed to the yeast's viability or to its content of nutrients, living and autoclaved yeasts were tested simultaneously. For this purpose, incubation trials were carried out using the long-term rumen simulation technique. Living and autoclaved yeasts were added to fermentation vessels at a concentration of 0.5 or 1.5 g/d. The addition of living and autoclaved yeasts stimulated microbial metabolism, with no major differences between the treatments. It was concluded that ruminal microbes digested the supplied yeast of S. boulardii as an additional substrate and that S. boulardii, at least in ruminants, is utilized as a prebiotic rather than as a probiotic agent.

Ammonia↗