Anorectal junction stenosis: diagnosis and management.

PURPOSE: The aim of this paper is to describe the management of 9 patients with anorectal junction stenosis and present the diagnostic features together with a single-stage surgical technique with reproducible results. MATERIALS AND METHODS: Nine patients with anorectal junction stenosis were seen over a period of 12 years. The children (aged from 2 months to 15 years) presented with constipation . Anorectal junction stenosis was diagnosed by rectal examination during which the tip of the finger was unable to pass beyond the stenosis in all cases. Contrast study showed the dilated rectosigmoid proximal to the stenosis. RESULTS: Six patients (who did not have significant rectal dilation) underwent single-stage surgery by posterior Y-V plasty which was curative. Two patients were operated through the posterior sagittal route with a covering colostomy done during the same session; the oldest child required resection of the megasigmoid with abdominoperineal pull-through. None of the six patients operated with Y-V plasty experienced any complications. One of the patients operated via the posterior sagittal route had a leak from the anorectal anastomosis requiring revision. The follow-up ranged from 6 months to 12 years. All patients are alive and well and there was no recurrence of stenosis in any case. CONCLUSION: Anorectal junction stenosis is a rare anorectal anomaly easily diagnosed by digital rectal examination. Single-stage surgery by posterior Y-V plasty is effective in curing the majority of these patients if significant rectosigmoid dilation is not present.

In vitro assessment of the cytotoxicity of nisin, pediocin, and selected colicins on simian virus 40-transfected human colon and Vero monkey kidney cells with trypan blue staining viability assays.

Gram-positive bacterial bacteriocins (nisin and pediocin) and gram-negative bacterial bacteriocins (colicins [Col] E1, E3, E6, E7, and K) were evaluated for cytotoxicity against cultured simian virus 40-transfected human colon (SV40-HC) and Vero monkey kidney (Vero) cells. Bacteriocin-treated cells were assessed for viability by trypan blue staining. Monolayers of SV40-HC and Vero cells were cultured in tissue culture plates (35 degrees C, 10% CO2 in humidified air) with the use of Dulbecco's modified Eagle's medium supplemented with 10% (vol/vol) calf serum. Actively growing cells in the log phase (ca. 10(4) cells per ml) were treated with individual partially purified bacteriocin preparations at 170, 350, and 700 activity units per ml. Duplicate culture plates for each bacteriocin treatment and untreated controls were withdrawn after 16, 32, and 48 h of incubation. Cells were dissociated with trypsin and treated with trypan blue and were then counted in a hemocytometer with the use of a phase-contrast microscope. Viability assays indicated dose-dependent toxicity for some bacteriocins. Nisin, pediocin, and Col E6 were the most cytotoxic bacteriocins; SV40-HC cells demonstrated greater sensitivity than Vero cells did. Some bacteriocins can be toxic to mammalian cells; therefore, bacteriocins intended for use as biopreservatives must be evaluated for toxicity to mammalian cells and for other toxicities. Col E1, Col E3, Col E7, and Col K demonstrated little toxicity at the activities tested, indicating that they are safe and thus have potential for use as food biopreservatives.

Study of the mechanism of the interaction of vinyl chloride with water by reversed-flow gas chromatography.

The investigation of the mechanism for the interaction of vinyl chloride (VC) with liquid foods is of great significance in food science. In the present work the model system VC-water was studied by using the relatively new technique of reversed-flow gas chromatography. Using suitable mathematical analysis the following physicochemical quantities were determined: (a) diffusion coefficients of VC into water, (b) overall mass transfer coefficients of VC in the water, and in the carrier gas nitrogen, (c) partition coefficients of VC between water and nitrogen, and (d) constants of Henry's law for the adsorption of VC by water. From the variation of the above parameters with temperature, and the stirring rate of the water, useful conclusions concerning the mechanism for the VC-water interaction were extracted. The experimental results for the transfer of VC into the bulk water suggest (i) a mechanism consisted of a fast equilibrium step between the VC in the gas phase and in the interface, followed by a slow adsorption of VC into the bulk of water, which is the rate-determining step, when the water is quiescent, and (ii) a mechanism consisted of a slow diffusion of VC from the gas phase to the interface, which is the rate-determining step, followed by a fast equilibrium step between the VC in the interface and in the water bulk, when the water is stirred.

Flux of gases across the air-water interface studied by reversed-flow gas chromatography.

In the present work the reversed-flow gas chromatographic technique was applied for the study of flux of gases across the air-water interface. The model system was vinyl chloride-water, which is of great significance in food and environmental chemistry. Using suitable mathematical analysis, equations were derived by means of which the following physicochemical quantities were calculated: diffusion coefficient of vinyl chloride (VC) into water, partition coefficient of VC between the water (at the interface and the bulk) and the carrier gas nitrogen, overall mass transfer coefficients of VC in the gas (nitrogen) and the liquid (water), gas and liquid film transfer coefficients of VC, gas and liquid phase resistances for the transfer of VC into the water, and finally the thickness of the stagnant film in the liquid phase, according to the two-film theory of Whitman. From the variation of the above parameters with temperature, as well as the volume and the free surface area of the water, useful conclusions concerning the mechanism for the transfer of VC into water were extracted. These are discussed in comparison with the same parameters calculated from empirical equations or determined experimentally by other techniques.

Solution of 1H NMR structure of the heme cavity in the oxygen-avid myoglobin from the trematode Paramphistomum epiclitum.

A two-dimensional 1H NMR study has been carried out on the heme cavity of the extreme oxygen-avid and autoxidation-resistant oxy-myoglobin complex from the trematode Paramphistomum epiclitum, and the residues were identified which potentially provide hydrogen bond stabilization for the bound oxygen. Complete assignment of the heme core resonances allows the identification of 10 key heme pocket residues, 4 Phe, 4 Tyr, and 2 upfield ring current aliphatic side chains. Based solely on the conserved myoglobin folding topology that places the E helix-heme crossover and the completely conserved Phe(CD1)-heme contact at opposing meso positions, the heme orientation in the cavity and the E helix alignment were unambiguously established that place Tyr66 at position E7. Moreover, all eight aromatic and the two aliphatic side chains were shown to occupy the positions in the heme cavity predicted by amino acid sequence alignment with globins of known tertiary structure. The dipolar contacts for the Tyr32(B10) and Tyr66(E7) rings indicate that both residues are oriented into the heme cavity, which is unprecedented in globins. The ring hydroxyl protons for both Tyr are close to each other and in a position to provide hydrogen bonds to the coordinated oxygen, as supported by strong retardation of their exchange rate with bulk solvent. A more crowded and compact structure increases the dynamic stability of the distal pocket and may contribute to the autoxidation resistance of this myoglobin.

Structural and functional comparison of trematode haemoglobins.

Haemoglobins from three different species of trematodes (Gastrothylax crumenifer, Paramphistomum epiclitum and Isoparorchis hypselobagri) and their respective hosts were digested with papain. Peptides so produced were separated on 10-15% SDS-PAGGE. The probability of coincidence of the peptides was analysed statistically. Oxygen affinity curves of trematode haemoglobins were found to be a rectangular hyperbolic. The Hill coefficient values of all the trematode haemoglobins were found to be 1. The P50 values, at 25 degrees C in 0.2M phosphate buffer pH 7.4, ranged from 0.8 to 1.6 mmHg.

Purification and properties of the hemoglobins of the platyhelminth Isoparorchis hypselobagri (Trematoda: Isoparorchidae) and its host Wallagu attu (catfish).

1. The hemoglobins of the trematode Isoparorchis hypselobagri and of its host Wallagu attu (catfish) were isolated and purified. 2. SDS-polyacrylamide gel electrophoresis showed both to consist of single, 15-17 kDa chains, having different electrophoretic mobilities. 3. Isoelectric focusing showed the trematode hemoglobin to be homogeneous with a pI of 4.2 and the host hemoglobin to consist of several components. 4. Gel filtration of freshly prepared trematode hemoglobin revealed one peak corresponding to M(r) approximately 17 kDa; gel filtration of a preparation which had been stored for 2-3 months demonstrated the presence of two peaks, whose elution volumes corresponded to M(r) of ca 35 and 17 kDa, respectively. 5. Reversed-phase chromatography of carboxymethylated 35 and 17 kDa peaks on a C8 column, gave a single peak a and two peaks b and c, respectively. 6. Edman degradation of peaks a, b and c obtained provided identical sequences of 27 amino acid residues for peaks a and c and another sequence differing at 10 of the 27 positions, for peak b. Edman degradation of the freshly prepared Isoparorchis hemoglobin provided the first 15 amino acid residues found for peaks a and c. The host hemoglobin gave an N-terminal sequence completely different from the trematode sequences. 7. Since gel filtration of the 35 and 17 kDa peaks showed no sign of an interconversion equilibrium, it appears that the 35 kDa peak and peak a represent a disulfide-bonded dimer of a monomer globin chain which shares the 27 N-terminal residues with chain c, the major monomer globin component of the 17 kDa peak.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparative spectral, electrophoretic and isoelectric properties of trematode haemoglobins.

A comparative study of the spectral, electrophoretic and isoelectric properties of the haemoglobins of three trematodes, Paramphistomum epiclitum, Gigantocotyle explanatum and Gastrothylax crumenifer was carried out. A high absorption in the beta band region indicates that trematode haemoglobins have high oxygen affinities. Electrophoretic mobilities of all trematode and their host haemoglobins were different. The isoelectric points of trematode haemoglobins were found to focus in the acidic range except that of G. crumenifer haemoglobin I, which focused at an alkaline pH.

Comparison of the hemoglobins of the platyhelminths Gastrothylax crumenifer and Paramphistomum epiclitum (Trematoda: Paramphistomatidae).

1. Gastrothylax crumenifer and Paramphistomum epiclitum parasitize the water buffalo Bubalus bubalis. 2. Gastrothylas hemoglobin consisted of two fractions of ca 30,000 and ca 18,000 by gel filtration. SDS-electrophoresis showed both to be single, ca 15,000 chains. 3. Paramphistomum hemoglobin was ca 16,000 by both gel filtration and SDS-electrophoresis. 4. Reversed-phase chromatography of carboxymethylated trematode and buffalo globins gave single peaks and two peaks, respectively. Although Paramphistomum hemoglobin provided and N-terminal sequence, Gastrothylax hemoglobin did not, suggesting blocked N-terminals. The buffalo sequences were found to be identical to the sequences of the alpha and beta chains of bovine hemoglobin. 5. Although Paramphistomum hemoglobin consists of only one chain, Gastrothylax hemoglobin consists either of one chain which aggregates to a dimer or of two different chains, only one of which aggregates to a dimer.

In vitro activation assays with hepatic S9 preparation from wild and laboratory reared woodchucks in the Salmonella mutagenicity test.

The Salmonella mutagenicity assay was utilized to compare hepatic S9 fractions derived from wild and laboratory reared woodchucks (Marmota monax). Two promutagens, 7,12-dimethylbenz[a]anthracence (DMBA) and 2-amino-fluorene (AF) were tested at 5 concentrations with the tester strains TA98 and TA100, against 2 levels of S9 fraction. AF produced similar number of revertants with the S9 fraction from wild and laboratory-reared animals. DMBA produced 2-4 times more revertant colonies at 50 microliter S9/plate with wild woodchuck S9 than with S9 from the laboratory-reared animals with both tester strains. It was concluded that natural inducers in the wild woodchuck diet may have contributed to the increased reversion frequency over laboratory reared woodchucks. Dose-response parameters for the activation of DMBA by S9 fraction from woodchucks and rats were compared with TA100. Woodchuck S9 had 3-40 more revertants/nmol and a 100-fold lower threshold of response than S9 from Aroclor 1254-induced rats.

Mutagenicity of chloroaniline/lignin metabolites in the Salmonella/microsome assay.

Chloroanilines are constituents of many agrochemicals and have been found to be metabolized to succinic acid conjugates, e.g., succinamides and succinimides. The mutagenic potential of five chloroanilines and their succinamides and succinimide derivatives have been tested with two strains of Salmonella typhimurium (TA98 and TA100) with and without rat hepatic microsomal fraction. None of the compounds produced a dose response effect with a two-fold increase in revertants indicating that these compounds are not mutagens or promutagens in these assays.

Mutagenic potency of some conjugated nitroaromatic compounds and its relationship to structure.

The mutagenicities of 12 conjugated non-fused nitroaromatic compounds and 1 amino analogue were determined in strains TA100 and TA98 of Salmonella typhimurium. Reversions by p-nitroaromatics increased in the order of the acetophenone, benzaldehyde, styrene, chalcone, cinnamic acid and stilbene indicating the importance for mutagenic potency of extended conjugation to the p-nitrophenyl substituent. Highest mutagenicity was found with alpha-substituted 4-nitrostyryl derivatives of which the phenyl derivative (31 revertants per nmole in TA100) was the most active. Generally, the TA100 strain was more sensitive than TA98 to these mutagens and S9 treatment was unnecessary for activity, although 4-nitrochalcone required S9 activation. Para-nitro isomers of the cinnamic acids and chalcones were much more active than the corresponding ortho and meta isomers. The 4-aminocinnamic acid analogue was inactive suggesting that complete reduction in Salmonella of 4-nitrocinnamic acid to an active amino derivative is not response for the high mutagenicity of the former. Mutagenicity of these p-nitrostyryl compounds may be explained by the covalent interaction of the electrophilic benzylic carbon with Salmonella DNA.

Residues and mutagenicity of captan applied to apple trees and potential human exposure.

The fungicide captan (cis-N-((trichloromethyl)thio) 4-cyclo-hexene-1,2-dicarboximide) was applied at the rate of 2.4 g/l to apple trees (c.v. Golden Delicious) individually or as part of a standard treatment program where it was applied eight times during the growing season together with several pesticides. Leaf samples (100 discs of 2.2 cm diameter) were collected from treated and control trees before treatment and at 0, 1, 3, 7, 14, 28, 56, 90 and 112 days after treatment. Fruit samples were taken at mid-season (56 days) and at harvest (112 days). The objective of this study was to determine the captan residue and mutagenicity of leaf and fruit extracts to ascertain the potential health hazard to agricultural workers in these orchards. Surface residues were extracted from leaves and fruits with methylene chloride. These extracts were subsequently analyzed for captan by gas-liquid chromatography (GLC) utilizing an electron-capture detector, and for mutagenicity with two strains (TA98 and TA100) of Salmonella typhimurium, with and without microsomal enzyme activation. Positive mutagenic effects were observed with strain TA100 at 0-14 days post spray, even with extracts from one leaf disc's surface (3.8 cm2) of the single treatment. Captan residues in these samples indicated a decline from 9.3 micrograms/cm2 at 0 days to 0.80 micrograms/cm2 at 14 days and a trace after 112 days. With the standard treatment, in which captan was incorporated eight times in the program starting at the 7-day interval, leaf extracts showed mutagenic activity at 7, 14, 28 and 90 days. Captan residues at these intervals were 11.4, 5.0, 4.1 and 3.4 micrograms/cm2, respectively. Fruit sample extracts of the standard spray were mutagenic to the tester strains TA100 and TA98 both at mid-season and at harvest. Residues of captan on fruits declined from 10.4 micrograms/cm2 at mid-season to 1.1 micrograms/cm2 at harvest. No mutagenic activity was detected with extracts from fruit samples from the single captan application.

Screening pesticides for their ability to damage bacterial DNA.

Twenty-six pesticides and pesticide degradation products were screened (125 micrograms - 2000 micrograms) for their ability to induce unrepairable damage to bacterial DNA. Three repair test systems were utilized in this study, the Salmonella typhimurium (TA1538/TA1978), the E. coli K-12 (Pol A1+/Pol1-) and the E. coli WP2 (WP2, WP2uvrA, WP67, CM611 and CM571). Aldicarb (1000 micrograms), benomyl (250 micrograms), 2-aminobenzimidazole (2000 micrograms), captan (125 micrograms), fenazalor (500 micrograms), 5,6-dichloro-2-trifluoromethylbenzimidazole (NC-2983) (250 micrograms), isothymol (250 micrograms), maleic hydrazide (1000 micrograms), pentachloronitrobenzene (1000 micrograms) were DNA-damaging to one or more bacterial test systems. Isothymol and NC-2983 affected all three test systems. Chlorinated hydrocarbon insecticides, some being recognized as carcinogens, did not produce a zone of inhibition in any of the tester strains possibly due to their poor solubility and diffusion in the agar overlay. It was concluded that these tests can be performed along with bacterial reversion tests to complement each other as short-term screening tests for potential carcinogens and mutagens.

Toxic and protective constituents in pet foods.

An analytical survey of mutagens, nitrosamines, polychlorinated biphenyls, toxic elements, and gamma-emission, as well as the toxicologically protective constituents zinc, selenium, and vitamin C, in 48 pet foods was conducted. Aside from high concentrations of fluoride and iodide in some samples and the expectedly higher concentrations of mercury and selenium in certain cat foods containing fish, the samples were notably free of the other toxic constituents. Direct-acting and promutagens and nitrosamines were not detectable in any of the samples. gamma-Emission was very low in all of the foods. Polychlorinated biphenyls were only detected in one cat food.

Evaluation of beta-naphthoxyacetic acid for mutagenic activity in the Salmonella/mammalian microsome assay.

Beta-naphthoxyacetic acid (BNOA) is used as a plant growth regulator on tomatoes and strawberries. It is the active ingredient in Blossom-Set and Berry-Set, two plant hormone sprays for fruit-set. The mutagenic activity of BNOA was evaluated in four strains of Salmonella typhimurium (TA97, TA98, TA100 and TA1535) in the presence and absence of liver microsomal and cytosolic enzymes derived from Aroclor induced rats. BNOA did not produce any significant increase (p less than 0.05) in the reversion of any of the four tester strains in the standard plate incorporation assay. Results of the agar overlay toxicity tests indicates that the chemical shows toxic effects at concentrations above 500 micrograms/plate. It was concluded that under the conditions of these tests, BNOA did not exhibit any mutagenic activity.

Structure-mutagenicity relationships of chalcones and their oxides in the Salmonella assay.

31 p-monosubstituted chalcones (E-1, 3-diphenylpropene-1-one) and the corresponding oxides (E-1-benzoyl-2-phenyloxirane) were tested for mutagenic activity on two strains of Salmonella typhimurium (TA98 and TA100) with and without rat liver microsomal and cytosolic enzymes. Highest mutagenicity (3.0 revertants/nmole in either strain) was seen with the 4-nitrochalcone, especially after S9 activation. Epoxidation, in general, increased the mutagenic activity of the respective chalcone. Benzoyl (4') substituted chalcones and their oxides with an electron-withdrawing substituent (e.g., nitro, fluoro) usually had higher activity than their phenyl (4) substituted counterparts, whereas the converse was the case with electron-donating substituents (e.g., acetamido, methoxy). Further multiple factorial analysis revealed that increasing hydrophilicity as indicated by the Hansch pi parameter, and resonance electronic contributions were more important than other factors including steric terms in explaining the mutagenicity of these compounds. Mutagenic effects of some chalcone oxides, particularly the 4-methoxy derivative, were markedly decreased by S9 treatment. The consequence of the weak-to-moderate mutagenicity of these compounds to dietary intake of hydroxylated and methoxylated chalcones is discussed.

Comparative mutagenicity tests in the Salmonella/microsome assay with rat and woodchuck S9 preparations.

The Salmonella mutagenicity assay was utilized to compare the hepatic S9 fractions from untreated and 3-methylcholanthrene (MC) induced woodchucks with Aroclor 1254 induced rats. Three known promutagens, benzo[a]pyrene (BP), 7,12-dimethylbenz[a]anthracene (DMBA), and 2-aminofluorene (AF) were tested at 5 concentrations with the strain TA100 against 3 levels of S9 fraction. Both woodchuck S9 fractions were as effective as the rat S9 in activating BP and both were more effective than the rat S9 in activating DMBA. Untreated woodchuck S9 was also as effective as rat S9 in activating AF. The protein content of the S9 fraction did not differ significantly between rats and woodchucks, but the P-450 content of the rat S9 was approximately 3.5 times that of woodchuck.

Mutagenicity tests with gallic and tannic acid in the Salmonella/mammalian microsome assay.

Gallic acid, tannic acid mixture and a purified fraction of tannic acid were evaluated for possible mutagenic activity in three strains of Salmonella typhimurium, TA98, TA100, and TA1535. These chemicals were not mutagenic either before or after activation with rat and woodchuck microsomal and cytosolic enzymes. However, tannic acid mixture and tannic acid fraction both gave a significantly (p = 0.05) dose-related reduction in the number of the revertant colonies, compared to the normal spontaneous revertants with no apparent toxic effects in the background lawn. With an agar diffusion assay, the chemicals exhibited toxic effects at 5000 micrograms/disc.