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Complete amino acid analysis in hydrolysates of foods and feces by liquid chromatography of precolumn phenylisothiocyanate derivatives.

The amino acid analysis method using precolumn phenylisothiocyanate (PITC) derivatization and liquid chromatography was modified for accurate determination of methionine (as methionine sulfone), cysteine/cystine (as cysteic acid), and all other amino acids, except tryptophan, in hydrolyzed samples of foods and feces. A simple liquid chromatographic method (requiring no derivatization) for the determination of tryptophan in alkaline hydrolysates of foods and feces was also developed. Separation of all amino acids by liquid chromatography was completed in 12 min compared with 60-90 min by ion-exchange chromatography. Variation expressed as coefficients of variation (CV) for the determination of most amino acids in the food and feces samples was not more than 4%, which compared favorably with the reproducibility of ion-exchange methods. Data for amino acids and recoveries of amino acid nitrogen obtained by liquid chromatographic methods were also similar to those obtained by conventional ion-exchange procedures.

Amino Acids↗

Natural infection with Eimeria leuckarti: prevalence of oocysts in feces of horse foals on several farms in Kentucky during 1986.

During 1986, fecal specimens were collected 1 or more times from each of 164 horse foals (158 Thoroughbred and 6 mixed light horse type), ranging in age from 0 to 252 days, on 13 farms in central Kentucky. To detect natural infection with Eimeria leuckarti, feces were examined for oocysts. Oocysts were found in 67 (41%) of the foals on 11 (85%) of the farms. The earliest age at which oocysts were first detected was 15 days (1 foal); the latest age was 123 days (1 foal). The mean age for the first appearance of oocysts in the feces of the 67 foals positive for E leuckarti was approximately 70 days. Age of the oldest foal that passed oocysts was 185 days. The longest oocyst shedding period was about 4 months, although oocyst-positive feces usually were found only sporadically during this period.

Age Factors↗

A survey of the aerobic bacteria in the feces of captive raptors.

Feces of 47 captive raptors belonging to the order Falconiformes or Strigiformes were cultured for bacteria. Gram-negative bacteria, which were cultured from the feces of 45 of the 47 raptors, were the most common isolates. A wide variety of species were identified, including a newly described genus (Moellerella wisconsensis), two newly described species (Escherichia fergusonii and Proteus penneri), and a member of a newly described enteric group (CDC Enteric group 41). Additional organisms identified that have not been reported in previous bacteriological surveys of raptors were Salmonella heidelberg, Salmonella braenderup, Morganella morganii, Yersinia ruckeri, Serratia spp., and Kluyvera sp. Escherichia coli, isolated from the feces of 42 of the 47 raptors, was the most frequently recovered. Streptococcus faecalis, the second most common isolate, was cultured from 30 birds. Several differences were observed between fecal bacteria isolated from raptors fed commercially prepared chicken and those isolated from raptors not fed chicken. The most obvious difference was that birds fed chicken had more varied gram-negative bacterial species and in greater numbers per fecal sample. The potential for the isolated bacteria from raptors as pathogens in humans and avian species is discussed.

Animals↗

An evaluation of alkaline peptone water for enrichment of Vibrio cholerae in feces.

A comparison was made to determine the sensitivity of direct inoculation of thiosulfate citrate bile salts agar (TCBS) and alkaline peptone water (APW) enrichment prior to direct inoculation of TCBS to culture Vibrio cholerae from feces of patients with gastroenteritis. V. cholerae was isolated from 611 feces specimens. Of those, 535 were isolated in TCBS and APW-TCBS, 15 in only TCBS and 61 in only APW-TCBS. V. parahemolyticus (21) and non-agglutinating vibrios (11) were also isolated but more often in direct inoculated TCBS than APW-TCBS cultures. Maximum isolation sensitivity of V. cholerae and V. parahemolyticus from feces is obtained by both direct inoculation of TCBS and enrichment in APW prior to TCBS inoculation.

Feces↗

Characterization of trisubstituted cholanoic acids in human feces.

The trisubstituted cholanoic acids in human feces have been studied by gas chromatography-mass spectrometry. The following bile acids have been identified: 3 Beta,7alpha,12alpha-trihydroxy-, 3 Beta,7 Beta,12alpha-trihydroxy-, 3alpha,7alpha-dihydroxy-12-keto-5 Beta-cholanoic acids and 3alpha,7alpha,12alpha-trihydroxy-5alpha-cholanoic acid. The presence in human feces of 3alpha,7alpha,12alpha-trihydroxy-, 3alpha,7,12alpha-trihydroxy-, and 3alpha,12alpha-dihydroxy-7-keto-5 Beta-cholanoic acids has been confirmed. The composition of bile acids in human feces is summarized and possible metabolic interrelationships suggested.

Bile Acids and Salts↗

Identification of mono- and dihydroxy bile acids in human feces by gas-liquid chromatography and mass spectrometry.

The mono- and disubstituted cholanoic acids present in human feces have been investigated. Extracts of feces were fractionated on silicic acid column and individual bile acids were isolated by preparative thin-layer chromatography. The isolated compounds were studied by gas-liquid chromatography of the methyl esters, partial trimethylsilyl ethers, oxidation products, and trifluoroacetates. The probable structures deduced were confirmed by gas chromatography-mass spectrometry and by comparisons with authentic compounds. The following derivatives of 5 Beta-cholanoic acid not previously isolated from human feces were identified: 3,12-diketo, 3-keto-12alpha-hydroxy, 3alpha,12 Beta-dihydroxy, 3 Beta,12 Beta-dihydroxy, 3-keto-7alpha-hydroxy, 3alpha-hydroxy-7-keto, 3 Beta,7alpha-dihydroxy, 3alpha,7alpha-dihydroxy, and 3alpha,7 Beta-dihydroxy. The presence of 3-keto-, 3 Beta-hydroxy-, 3alpha-hydroxy-, 3 Beta-hydroxy-12-keto-, 3alpha-hydroxy-12-keto-, 3 Beta,12alpha-dihydroxy-, and 3alpha,12alpha-dihydroxy-5 Beta-cholanoic acids was confirmed. Evidence was obtained for the presence of two bile acids having at least one hydroxyl group at a carbon atom other than C(3), C(7), or C(12).

Bile Acids and Salts↗

Immunochemical differences among carcinoembryonic antigen in tumor tissues and related antigens in meconium and adult feces.

We have isolated carcinoembryonic antigen (CEA)-related antigens from meconium and compared them with those in adult feces. Two CEA-related antigens were detected in meconium [nonspecific cross-reacting antigen 2 (NCA-2) and meconium nonspecific cross-reacting antigen] while four CEA-related antigens were found in adult feces [normal fecal antigen 1, normal fecal antigen 2 (NFA-2), normal fecal cross-reacting antigen, and fecal nonspecific cross-reacting antigen, respectively]. By conventional anti-CEA antisera, NCA-2 in meconium, NFA-2 in adult feces, and CEA in tumor tissues were indistinguishable from each other, but they could be distinguished by specific antibody preparations against a determinant unique to CEA (CEA-distinctive determinant) or to NFA-2 (NFA-2-distinctive determinant). Neither the CEA-distinctive determinant nor the NFA-2-distinctive determinant was detected on the NCA-2 molecule. No antigenic determinants unique to NCA-2 have been detected with the anti-NCA-2 antisera which we have prepared thus far. The molecular weight of purified NCA-2 was estimated to be 150,000 to 170,000 as compared to 160,000 to 170,000 for NFA-2 and 170,000 to 180,000 for CEA. NCA-2 had amino acid and carbohydrate compositions similar to those of CEA and NFA-2. All NFA-2 preparations and about one-half of the CEA preparations were sensitive to Pronase E digestion, which released two antigen fragments from these molecules, but NCA-2-preparations were resistant to such digestion.

Amino Acids↗

Escherichia coli heat-stable enterotoxin in feces and intestines of calves with diarrhea.

Two experiments were conducted to evaluate detection of Escherichia coli heat-stable enterotoxin (ST) in the feces of calves as a method for implicating E coli in neonatal calf diarrhea. The first experiment evaluated the use of the infant mouse test for detection of ST in the feces of calves with naturally occurring diarrhea. Simultaneous identification of bovine enteropathogenic strains of E coli (EEC) and of other infective agents implicated in neonatal calf diarrhea was attempted in these samples. The ST was detected with certainty in only 7 of 41 samples from calves less than or equal to 3 weeks old. Enteropathogenic E coli, however, was detected in 27 samples. In 23 of these 27 samples, EEC was the only recognizable diarrheagenic agent. In a small percentage of the samples, Salmonella, rotavirus, coronavirus, and cryptosporidium were recognized alone, in combination with each other, or with EEC. In the second experiment, 6 calves were fed colostrum from cows inoculated with the bovine EEC strain B44; 6 were given colostrum from cows vaccinated with non-EEC strain 28F, and 4 were given milk from nonvaccinated heifers. Two of the calves that were given colostrum from cows inoculated with strain B44 were challenge exposed with the non-EEC strain 28F. The remaining calves were challenge exposed with the EEc strain B44. Fecal samples were taken from these calves at intervals and were examined for the presence of ST and of the challenge-exposure organism. The ST was detected in approximately one half of the fecal samples obtained, and it was most often detected in the early stages of the induced diarrhea. Calves were observed to shed the challenge-exposure EEC strain for long periods in the absence of diarrhea or detectable amounts of ST in the feces. The ST was detectable in fecal samples when the diarrhea was severe and when the dry matter content of the fecal samples was low.

Animals↗

Enzyme-linked immunosorbent assay, using monoclonal antibody, to detect enterotoxic Escherichia coli K99 antigen in feces of dairy calves.

A modified, double-antibody, enzyme-linked immunosorbent assay (ELISA) was developed to detect the K99 pilus antigen of enterotoxic Escherichia coli (ETEC) in feces of calves. Extremely high positive to negative ratios (greater than 200) were obtained by using monoclonal antisera as the primary antibody. Strong positive reactions were obtained with strains of E coli known to produce the K99 antigen; however, non-enteropathogenic E coli (strains not producing the K99 antigen), Salmonella, Proteus, Klebsiella, Pseudomonas, Staphylococcus, Streptococcus, and rotavirus produced negative results. Seventy-five fecal samples, 8 from healthy calves and 67 from calves with neonatal calf diarrhea were examined with the K99 ELISA for the presence of ETEC. Rotavirus test and fecal culture results were available on feces from calves with diarrhea and were used with the K99 ELISA results to determine the specific cause of the disease. Enterotoxic E coli was the predominant agent detected in the feces of 29 diarrheal calves less than 5 days of age. Mixed infections of rotavirus and ETEC were also common in these calves, but rotavirus infections alone were not detected. In 38 calves greater than or equal to 5 days, rotavirus was detected without ETEC. Of these calves, only 2 produced positive tests with the K99 ELISA. Salmonella sp and Proteus sp were detected from 5 of 67 calves with diarrhea.

Agglutination Tests↗

In vitro fermentation of feces from normal and chronically diarrheal horses.

Feces from 13 healthy horses and 8 horses with chronic diarrhea were subjected to an in vetro fermentation procedure that had been developed for rumen fluid. Fermentations were conducted over 6 hours in a closed system, with and without an essential amino acid (EAA) mixture being added to the basic starch-buffer medium. The addition of EAA caused no significant difference in results of fermentation of feces from healthy horses. For diarrheic animals, there was a significant (P less than 0.01) increase in gas and total volatile fatty acids production whether EAA were present or not, and alpha-amino nitrogen was utilized in significantly (P less than 0.01) greater amounts only if EAA was present. Fermentations were repeated on feces from five of the eight diarrheal horses after they had been treated with oral iodochlorhydroxyquin for 1 week, and had shown desirable clinical response. A significant difference was not shown between pre- and posttreatment fermentations, except for decreased butyrate production. The results are consistent with the hypothesis that chronic equine diarrhea is primarily a colonic disease and indicates that colonic maldigestion may, in part, be responsible for excess fecal water.

Amino Acids, Essential↗

Quantitative culture of Rhodococcus equi from the feces of horse.

The selective isolation of R. equi in NANC medium was made by quantitative culture of the organism in the feces. R. equi was observed in the feces of all the mares and foals investigated. The mean viable count of R. equi organisms in 1 gram of feces was 8.42 X 10(2) in the mares and 7.57 X 10(2) in the foals, and its 95% confidence limits were in a range of 6.48 X 10(2) to 1.09 X 10(3) and 4.19 X 10(2) to 1.37 X 10(3), respectively. This result indicates that R. equi is a member of the normal intestinal flora in the horse.

Animals↗

[Effect of gum guar diet on feces microbiology in rats].

Approximately 8--10 days after gum guar was added to the diet of rats, the total bacterial counts in their feces, in particular Clostridium and Lactobacillus, were increased, whereas the number of coliforms was reduced. In contrast to the long, slender, rod-shaped cells in the Gram-stained fecal smears of the control animals, the feces of the gum guar-fed animals showed a predominance of short, broad, Gram-positive rods. When gum guar was omitted from the diet of these animals, their feces could not be distinguished from those of control animals.

Animals↗

Short-chain fatty acids in the normal human feces.

The short-chain fatty acids ( SCFAs ) have been studied in the feces of 20 healthy subjects--10 methane excretors and 10 non-methane excretors. The analytical procedure included homogenization of fecal samples followed by vacuum distillation and subsequent gas chromatography. This method for analysis of fecal SCFAs showed recoveries of the individual acids from 90% to 109% and coefficients of variation for the inter-assay reproducibility from 6.0% to 19.7%, highest for those acids present in the smallest concentrations. There was no difference in the concentrations or relative compositions of SCFAs between methane-excreting subjects and non-methane-excreting subjects. The concentrations of SCFAs , given as mmol/kg feces (wet weight), were (median and range): total, 76.8 (27.9-187.7); acetic acid, 37.4 (12.8-103.4); propionic acid, 12.5 (4.5-27.8); i-butyric acid, 2.2 (0.7-3.8); n-butyric acid, 12.4 (4.0-53.0); i-valeric acid, 3.2 (0.8-5.9); n-valeric acid, 2.4 (0.6-3.8) and n-caproic acid, 0.5 (0.0-3.6). The study shows that the SCFAs are quantitatively the most important anions in the feces of healthy subjects. The pronounced individual variations in the concentrations of SCFAs are real biological variations and cannot be explained by methodological variations.

Adult↗

Non-volatile N-nitroso compounds in human feces.

A bacterial mutagen has been detected in ether extracts of freeze-dried feces of humans on Western diet. Several types of evidence suggest that nitroso compounds are present in human feces and the mutagen could be a nitroso compound. Ascorbic acid supplement in the diet reduces the levels of both the mutagens and nitroso compounds in the feces.

Ascorbic Acid↗

Chemical test for mammalian feces: collaborative study.

A chemical method was developed to determine the presence of mammalian feces in food. The method consisted of the colorimetric determination of alkaline phosphatase and involved the splitting of the phosphate radical and phenolphthalein diphosphate and the resultant color development of phenolphthalein in a basic medium. A collaborative study testing the feces of 22 animals resulted in a 95% correct identification of samples as mammalian feces. The method has been adopted official first action.

Alkaline Phosphatase↗

Slide coagglutination for Salmonella typhi antigens in broths inoculated with feces from typhoid fever patients.

Salmonella typhi antigens D, Vi and d were readily detected, by slide coagglutination, in mannitol selenite (MSB) and dulcitol selenite (DSB), Salmonella enrichment broths 4 hours after inoculation with feces from 60 patients with bacteriologically confirmed typhoid fever. Positive coagglutination also occurred using MSB and DSB inoculated with fecal specimens obtained from 16 patients from whom S. typhi was not cultured. Twelve of these later seroconverted to Salmonella O antigen. None of the MSB or DSB inoculated with feces from 50 healthy control subjects, gave a positive coagglutination test. The coagglutination method appears to have potential as a rapid test for the detection of antigens of S. typhi in MSB and DSB broths inoculated with feces from patients with suspected typhoid fever.

Agglutination Tests↗

Detection of K-ras mutations in DNAs isolated from feces of patients with colorectal tumors by mutant-allele-specific amplification (MASA).

K-ras mutations are found in approximately half of all colorectal tumors examined. To explore the possibility of detecting mutated K-ras rapidly and efficiently in DNAs isolated from fecal material, we applied the mutant allele specific amplification (MASA)-PCR method to DNA from feces of patients with colorectal tumors. Among 55 colorectal adenocarcinomas or adenomas examined, 19 were found to carry K-ras mutations in codons 12 or 13. We were able to PCR-amplify DNAs isolated from feces of 15 of these 19 patients, but in only three of the fecal samples, we were able to detect the K-ras mutations corresponding to tumor DNA by MASA and ethidiumbromide staining of the gel. The carcinomas in these three cases were more than 40 mm x 40 mm in size and located in the sigmoid colon or rectum. However, we identified the K-ras mutations in fecal DNAs of additional seven patients by MASA when the gels were blotted and probed with a radio-labeled oligonucleotide; the tumors in those patients had arisen in the distal half of the colon and the smallest of these tumors was only 7 mm x 5 mm. No K-ras mutations were detectable in feces of the remaining five cases, whose tumors were relatively small and/or located in the proximal region. The results suggested that the MASA-PCR system has potential for development as a simple, rapid and noninvasive method for diagnosing the presence of colorectal tumors that carry mutant K-ras alleles, particularly tumors located in the distal colon.

Adenocarcinoma↗

Anticoagulant effects of warfarin and kinetics of K vitamins in blood and feces.

Patients (40 cases) were treated with daily dosage of warfarin of 2-7 mg after being undergone artificial valve replacements. Twenty one days after administration of warfarin, we examined the patients for kinetics of K vitamins and vitamin K-dependent coagulation factors in blood, and intestinal flora in feces, as well as the relationship between K vitamins and coagulation activity. The following results were obtained. (1) In warfarin-administered patients (Group B), blood levels of vitamin K1 and menaquinone-7, a vitamin K2 homologue, were similar to those in non-warfarin-administered patients. Therefore, administration of warfarin did not significantly decreased the levels. (2) In patients selected randomly from Group B (Group C), the vitamin K1 level in feces was higher than that in non-warfarin-administered patients. The menaquinone-7 level in feces was similar to that in non-warfarin-administered patients. For the total counts of bacteria and the detection rate of vitamin K2-producing bacteria, there was no significant difference between Group C and non-warfarin-administered patients. (3) The above mentioned results of (1) and (2) suggest that it is important for development of anticoagulant effects by warfarin to inhibit conversion from vitamin K1 to reduced vitamin K1, as well as to inhibit the reducing process from vitamin K1-epoxide to vitamin K1. (4) Vitamin K1-epoxide, a metabolite of vitamin K1, appeared in blood after administration of warfarin; there was a lower correlation between the blood level of vitamin K1-epoxide and the warfarin dosage. Further, PIVKA-II appeared in blood after administration of warfarin; there was a inverse lower correlation between the level of PIVKA-II and HPT, and between PIVIKA-II and TT. In conclusion, it has been clarified that vitamin K1-epoxide and PIVKA-II are useful parameters to evaluate anticoagulant effect of warfarin.

Adult↗