Excited-state dipole moments of some hydroxycoumarin dyes using an efficient solvatochromic method based on the solvent polarity parameter, EN(T).

Excited-state dipole moments of some hydroxycoumarins, extensively used as laser dyes, have been determined using the solvatochromic method based on the microscopic solvent polarity parameter EN(T). Agreement between experimental and Austin model 1 (AM 1) calculated dipole moment changes has been found to be close in most of the cases. Our results are expected to be quite reliable in view of the fact that the correlation of the solvatochromic Stokes shifts is superior to that obtained using bulk solvent polarity functions. The dipole moments in the excited state, for all the molecules investigated, are higher than the corresponding values in the ground state. The increase in dipole moment upon excitation has been explained in terms of the nature of emitting state and resonance structure.

Delayed onset generalised dystonia after cyanide poisoning.

A 27 year old female developed delayed onset of persistent generalized dystonia following a suicidal attempt with potassium cyanide. Cranial CT scan showed bilateral putaminal hypodensities which were also seen on MRI scans to be hypointense on T1 and hyperintense on T2 weighted images. Multimodality evoked potentials were normal. An improvement was noted with levodopa.

Loss of calcium homeostasis leads to progressive phase of chlordecone-potentiated carbon tetrachloride hepatotoxicity.

Earlier work has shown increased hepatocellular free Ca2+ levels in rats receiving a single subtoxic dose of CCl4 after dietary pretreatment with nontoxic (10 ppm, 15 days) levels of chlordecone (CD), indicating a significant perturbation of Ca2+ homeostasis in the interactive toxicity of CD + CCl4 combination treatment. In the present study, the mitochondrial and microsomal ability to sequester Ca2+ as well as plasma membrane translocase activity was investigated, since it is known that cells maintain normal Ca2+ homeostasis by these mechanisms. Hepatic plasma membrane Ca(2+)-ATPase (high and low affinity components) as well as 45Ca uptake by mitochondria and microsomes was measured using a range of calcium concentrations in Ca(2+)-EGTA-buffered medium at different time points after a single ip administration of CCl4 (100 microliters/kg). Male Sprague-Dawley rats were maintained for 15 days either on a normal diet or on a diet containing 10 ppm CD prior to CCl4 injection. Hepatic plasma membranes, devoid of microsomal and mitochondrial contamination, were prepared using polyethyleneimine-coated beads. CD treatment alone did not significantly decrease the plasma membrane Ca(2+)-ATPase activity. Similarly, CCl4 treatment alone did not alter Ca(2+)-ATPase in hepatic plasma membranes at any concentration of free Ca2+ in assay medium employed in this study. The interactive combination treatment, however, resulted in significant, irreversible, and specific inhibition of the high affinity component of the hepatic plasma membrane Ca(2+)-ATPase at early time points. Low affinity Ca(2+)-ATPase was not affected with any treatment protocol. CD pretreatment alone significantly inhibited 45Ca uptake by mitochondria and microsomes when incubated at 10 microM and higher, concentrations much higher than normal cytosolic levels, but not at lower concentrations of Ca2+. CCl4 administration to both normal and CD-pretreated rats resulted in significant inhibition of microsomal and mitochondrial 45Ca uptake as early as 1 hr at all concentrations of free calcium. While the extent of inhibition was greater and irreversible after CD + CCl4 treatment, it was reversible after normal diet + CCl4 treatment. Phosphorylation of proteins was determined in order to investigate if the inhibition of microsomal 45Ca uptake during CD + CCl4 toxicity might be correlated to decreased phosphorylation of any particular protein involved in Ca2+ transport. SDS-polyacrylamide gel electrophoresis of microsomal protein revealed at least 30 Coomassie blue stainable bands. Of these, 6 bands were phosphorylated when microsomes were incubated with [32P]ATP.(ABSTRACT TRUNCATED AT 400 WORDS)

Effect of antimitotic agent colchicine on carbon tetrachloride toxicity.

A single administration of a subtoxic dose of CCl4 (100 microliters/kg, i.p.) is known to induce hepatocellular regeneration and tissue repair at 6 and 48 h in rats, permitting prompt recovery from the limited liver injury associated with that dose of CCl4. Substantial evidence has accumulated to indicate that the early-phase hepatocellular regeneration and tissue repair are critical for recovery from halomethane hepatotoxicity. The objective of these studies was to test this concept in an experimental framework, wherein a selective ablation of the early-phase cell division should result in prolongation of liver injury followed by recovery. The studies were designed to evaluate the influence of the antimitotic agent colchicine (1 mg/kg, i.p. in saline) on CCl4 toxicity. Colchicine was administered 2 h prior to CCl4 or corn oil injection. Toxicological end points and markers of hepatocellular regeneration were assessed at various time points (2, 6, 12, 24, 48 and 72 h) after the injection of CCl4 to male Sprague-Dawley rats. Hepatocellular injury was assessed through elevations of serum alanine and aspartate aminotransferase and by histopathological examination of the liver. Incorporation of 3H-thymidine in hepatocellular nuclear DNA and mitotic index were used as indices of hepatocellular regeneration. Hepatocellular regeneration stimulated by CCl4 at 2-6 h was blocked by colchicine as evidenced by the decreased 3H-thymidine incorporation and mitotic index,without any significant effect on the second phase of cell division at 48 h. Ablation of this early phase of tissue repair resulted in prolongation of CCl4 hepatoxicity.(ABSTRACT TRUNCATED AT 250 WORDS)

Colchicine antimitosis abolishes CCl4 autoprotection.

A subtoxic dose of CCl4 is known to destroy liver microsomal cytochrome P-450 and this is widely accepted as the mechanism of CCl4 autoprotection. Circumstantial evidence suggests that while cytochrome P-450 is significantly decreased, this mechanism alone cannot explain the phenomenon of autoprotection. Previous studies have established that hepatocellular regeneration is stimulated as early as 6 hr after the administration of a low dose of CCl4. If the early phase stimulation of hepatocellular regeneration by the protective dose is indeed the mechanism of autoprotection, then ablation of this early phase of tissue healing by colchicine should result in an abolishment of autoprotection. Present studies were conducted to test this conceptual premise. The protection afforded by a low dose of CCl4 (LCCl4, 100 microliter/kg, ip) on the toxic effects of a subsequently administered moderately toxic dose of CCl4 (HCCL4, 2.5 ml/kg, ip) was established in male Sprague-Dawley rats. The protective dose provided 100% protection, whereas only 62.5% survival was observed when the corn oil vehicle was administered instead of the protective dose of LCCl4. Colchicine administration (1 mg/kg, ip in saline) 2 hr prior to the injection of LCCl4 led to a complete loss of autoprotection resulting in 100% mortality in rats given the HCCl4. Earlier studies have established that colchicine selectively suppresses the early phase of hepatocellular regeneration at 6 hr without influencing the second phase at 36-48 hr. The consequence of colchicine antimitosis on the toxicological endpoints of liver injury was evaluated by serum enzyme elevations and by histopathological examination of the liver during a time course of 6, 24, 48, 72, and 96 hr after the administration of HCCl4. In the autoprotection regime, after only a transient and modest elevation of serum alanine and aspartate transaminases, complete recovery occurred by 96 hr. Hepatocellular necrosis was consistently lower compared to all other groups. Colchicine preadministration in the autoprotection regime resulted in significantly greater and progressive elevation of the serum enzymes and a correspondingly commensurate progression of hepatic lesion. Toxic effects of HCCl4 alone were more rapidly and maximally augmented by colchicine preadministration. The role of hepatocellular regeneration in autoprotection was evaluated by 3H-thymidine incorporation in hepatocellular nuclear DNA and by morphometric estimation of mitotic index. While HCCl4 alone resulted in some stimulation of 3H-thymidine incorporation and mitosis, the regenerative activity observed with prior LCCl4 administration was remarkably greater, particularly at 48 hr. Colchicine preadministration in either of these 2 protocols decisively obtunded the stimulated regenerative activities essentially abolishing the tissue healing mechanisms.(ABSTRACT TRUNCATED AT 400 WORDS)

Development of a method for concentration of rotavirus and its application to recovery of rotaviruses from estuarine waters.

As part of our studies on the ecology of human enteric viruses, an improved method for detection of rotaviruses in water was developed, and their presence in Galveston Bay was monitored. Samples (378 liters) of estuarine water adjusted to pH 3.5 and a final AlCl3 molarity of 0.001 were filtered through 25-cm pleated cartridge-type filters (Filterite Corp., Timonium, Md.) of 3.0- and 0.45-micron porosity. Adsorbed virus was eluted with 1 liter of 10% tryptose phosphate broth, pH 9.5. Primary eluates were reconcentrated to a final volume of 10 to 20 ml by a simple and rapid magnetic iron oxide adsorption and elution procedure. Two percent casein at pH 8.5 effectively eluted rotavirus from iron oxide. A total of 21 of 72 samples of water, suspended solids, fluffy sediments, and compact sediments collected in different seasons in Galveston Bay yielded rotaviruses. Recovery of rotaviruses varied from 119 to 1,000 PFU/378 liters of water, 1,200 PFU/1,000 g of compact sediment, 800 to 3,800 PFU/378 liters of fluffy sediment, and 1,800 to 4,980 PFU from suspended solids derived from 378 liters of water based on immunofluorescent foci counts on cover slip cultures of fetal monkey kidney cells.

Human viruses in sediments, sludges, and soils.

Recent studies have provided a greater understanding of the movement of viruses in the environment by their attachment to solids. These studies have focused on solids-associated viruses present in wastewater discharged into the ocean and on viruses in sludge and wastewater that may be retained in soil following their land disposal. Such ocean or land disposal of wastewater and sludge may result in a discharge of one or more of 120 human enteric virus pathogens including those causing poliomyelitis, viral hepatitis A and acute gastroenteritis.Solids-associated viruses in effluents discharged into coastal waters accumulate in bottom sediments, which may contain 10 to 10 000 more virus per unit volume than the overlying seawater. Solids-associated viruses resuspended by water turbulence may be transported from polluted to distant non-polluted recreational or shellfish-growing water. Transmission of viruses causing hepatitis or gastroenteritis may result from contact by bathers or swimmers with these viruses in recreational waters, or from ingestion of raw or improperly cooked shellfish in which the solids-associated virus had been bioaccumulated.The land disposal of sludge and wastewater has a potential of causing infections in farm workers, contamination of crops, pollution of raw potable water sources or infiltration of ground water. Viruses retained on soils can be released by rain water and may contaminate ground water through lateral and vertical movements.

Method for detecting viruses in aerosols.

A simple method with poliovirus as the model was developed for recovering human enteric viruses from aerosols. Filterite filters (pore size, 0.45 micron; Filterite Corp., Timonium, Md.) moistened with glycine buffer (pH 3.5) were used for adsorbing the aerosolized virus. No virus passed the filter, even with air flow rates of 100 liters/min. Virus recovery from the filter was achieved by rapid elution with 800 ml of glycine buffer, pH 10. The virus in the primary eluate was reconcentrated by adjusting the pH to 3.5, adding AlCl3 to 0.0005 M, collecting the virus on a 0.25-micron-pore Filerite disk (diameter, 25 mm) and and eluting with 6 ml of buffer, pH 10. With this method, virus could be detected regularly in aerosols produced by flushing when 3 X 10(8) PFU of poliovirus were present in the toilet bowl. Poliovirus-containing fecal material from two of four infants who had recently received oral polio vaccine also yielded virus in the aerosols when feces containing 2.4 X 10(7) to 4.5 X 10(7) PFU of virus had been added to the toilet bowl. Persons infected with a variety of natural enteric viruses are known to excrete this amount of virus in their daily stools.

Isolation of enteroviruses from water, suspended solids, and sediments from Galveston Bay: survival of poliovirus and rotavirus adsorbed to sediments.

The distribution and quantitation of enteroviruses among water, suspended solids, and compact sediments in a polluted estuary are described. Samples were collected sequentially from water, suspended solids, fluffy sediments (uppermost layer of bottom sediments), and compact sediment. A total of 103 samples were examined of which 27 (26%) were positive for virus. Polioviruses were recovered most often, followed by coxsackie B viruses and echoviruses 7 and 29. Virus was found most often attached to suspended solids: 72% of these samples were positive, whereas only 14% of water samples without solids yielded virus. Fluffy sediments yielded virus in 47% of the samples, whereas only 5% of compact bottom-sediment samples were positive. When associated with solids, poliovirus and rotavirus retained their infectious quality for 19 days. The same viruses remained infectious for only 9 days when freely suspended in seawater. Collection of suspended solids at ambient water pH appears to be very useful for the detection of virus; it has advantages over collecting and processing large volumes of water, with accompanying pH adjustment and salt addition for processing.

Round robin investigation of methods for the recovery of poliovirus from drinking water.

Six laboratories actively involved in water virology research participated in a methods evaluation study, conducted under the auspices of the American Society for Testing and Materials Committee on Viruses in the Aquatic Environment, Task Force on Drinking Water. Each participant was asked to examine the Viradel (virus adsorption-elution) method with cartridge-type Filterite filters for virus adsorption and organic flocculation and aluminum hydroxide-hydroextraction for reconcentration. Virus was adsorbed to filter media at pH 3.5 and eluted with either glycine buffer (pH 10.5) or beef extract-glycine (pHG 9.0). Considerable variation was noted in the quantity of virus recovered from four 100-liter samples of dechlorinated tapwater seeded with low (350 to 860 PFU) and high (1,837 to 4,689 PFU) doses of poliovirus type 1. To have a more uniform standard of comparison, all the test samples were reassayed in one laboratory, where titers were also determined for the virus seed. Test results of the Viradel-organic flocculation method indicated that the average percentage of virus recovery for low-input experiments was 66%, with a range of 8 to 20% in two laboratories, 49 to 63% in three laboratories, and 198% in one laboratory. For the high-input experiments, two laboratories reported recoveries of 6 to 12%, and four laboratories reported recoveries of 26 to 46%. For the Viradel aluminum hydroxide-hydroextraction procedure, two laboratories recovered 9 to 11%, whereas four obtained 17 to 34% for low-input experiments. For the high-input tests, two laboratories reported a recovery of 3 to 5%, and four recovered 11 to 18% of the seeded virus.(ABSTRACT TRUNCATED AT 250 WORDS)

Concentration of rotavirus and enteroviruses from blue crabs (Callinectes sapidus).

A simple method for concentration and detection of rotavirus and enteroviruses in the blue crab is described. Virus was separated from tissue homogenates at pH 9.5, concentrated by adsorption to protein precipitates at pH 3.5, and recovered by elution of precipitates at pH 9.2. Test samples of 12 to 15 ml were produced from an initial 100 g of crab tissues. Cat-floc precipitation was used to remove sample toxicity for cell cultures. Recovery effectiveness averaged 52% with poliovirus 1, echovirus 7, and coxsackievirus B5 and 23% with rotavirus SA11.

Reduction of cytotoxicity in virus concentrates from environmental samples.

The reduction of cytotoxicity in virus concentrates from environmental samples was accomplished by high-speed centrifugation and by filtration of the samples through positively charged depth filters.

Detection of viruses in drinking water by concentration on magnetic iron oxide.

Discharge of raw domestic wastes containing human enteric viruses into water courses, consumption of untreated water from canals, streams, and shallow wells in villages, and cross-contamination of water in the distribution system because of intermittent water supply in urban areas continue to cause widespread outbreaks of infectious hepatitis in India. To detect a low number of viruses in 50- to 100-liter samples of water, a method was developed with magnetic iron oxide as the virus adsorbent. Poliovirus-seeded dechlorinated tap water, adjusted to pH 3.0 and 0.0005 M AlCl3, was filtered through a 10-g bed of iron oxide sandwiched between two AP20 prefilter pads held in a 142-mm-diameter, stainless-steel holder. Virus was eluted from iron oxide by recirculating three times a 100-ml volume of 3% beef extract, pH 9.0. The eluate was reconcentrated to 5 ml by adjusting to pH 3, adding 1 g of iron oxide, stirring for 30 min, and eluting the readsorbed virus with 5 ml of beef extract, pH 9.0. Virus recovery varied from 60 to 80%. Using the above method, we took a survey of drinking water at three locations in Nagpur during 1976 and found the presence of virus in 7 of 50 samples. The quantity of virus recovered ranged from 1 to 7 plaque-forming units per 30 to 60 liters. Virus was detected in some samples even with residual chlorine. No coliforms were detected in the virus-positive samples.

Glass Wool-Hydroextraction Method for Recovery of Human Enteroviruses from Shellfish.

The Glass Wool-Hydroextraction Method was developed to analyze a number of foods for the presence of contaminating human enteroviruses. This method was modified to examine a variety of shellfish, including oysters and hard- and soft-shell clams. The method consistently recovered ca. 50% of viruses inoculated into shellfish at levels of ca. 10 virus units/100 g. In a multilaboratory study, the method successfully detected all but one of the eight test viruses, and the quantitative recoveries compared favorably with the control laboratory data.