Evaluation of a novel, rapid-acting, sterilizing solution at room temperature.

Test data are presented for a novel chemical germicide formulation capable of sterilizing reusable medical devices in 30 minutes at 20 degrees C in an open tray. The tests conducted with this rapid-acting sterilizing solution (RSS) included sporicidal, mycobactericidal, and virucidal studies performed in accordance with Association of Official Analytical Chemist International or Environmental Protection Agency published guidelines, by using RSS stressed for as long as 7 days. Sporicidal assays were performed at 20 degrees C with a 30-minute exposure time by using both Clostridium sporogenes and Bacillus subtilis spores dried on porcelain penicylinders or suture loops (n = 60 carriers per treatment). For comparison, identical carriers were exposed to a commercial glutaraldehyde-based sterilizing solution stressed to a maximum-use time of 14 days and exposed per manufacturer's requirements (10 hours at 25 degrees C). The RSS sterilized 100% of the carriers of both spore types. The glutaraldehyde solution demonstrated 100% sterilization of C sporogenes -treated carriers but had difficulty sterilizing B subtilis spore-laden carriers (ie, no sterilization of suture loops and only 57% sterilization of porcelain penicylinders). Similarly, Mycobacteria bovis and selected fungal and viral agents were exposed to stressed solution for 5 minutes or less at 20 degrees C. In each case, the resulting log decrease in viable microorganisms significantly supported a claim for rapid high-level disinfection. Based on these data, RSS demonstrates high-level disinfection in 5 minutes and sterilization in 30 minutes at 20 degrees C.

Development and evaluation of a new alcohol-based surgical hand scrub formulation with persistent antimicrobial characteristics and brushless application.

BACKGROUND: Since the introduction in the 1970s of surgical hand scrub formulations that contain 4% chlorhexidine gluconate (CHG), new surgical scrub formulations that have improved efficacy, persistence, or significantly improved use characteristics have not been forthcoming. In addition, the manufacturer's labeling for popular hand scrub products generally requires scrub times in excess of 6 minutes, whereas current practical needs call for products with substantially shorter scrub times. A new alcohol-based surgical scrub formulation, which has ingredients that provide emollient, surfactant, and antimicrobial persistence characteristics to complement the rapid and broad-spectrum antiseptic qualities of alcohol, has been developed in an effort to address these current practical needs. METHODS: The relative efficacy of a new alcohol-based surgical scrub formulation that contains ingredients that provide surfactant and antimicrobial persistence characteristics was compared with that of commercial 4% CHG and 7.5% povidone iodine (PVPI) formulations with use of human subjects. Hand antimicrobial count sampling was performed by using standardized "glove juice" methodology. RESULTS: The efficacy and persistence results of the new formulation showed statistically significant improvement over both CHG and PVPI at a substantially lessened scrub time (3 minutes). In addition, use of the new formulation without a scrub brush produced results statistically similar to 3-minute applications with either a brush or a sponge. CONCLUSIONS: The new alcohol-based formulation demonstrates promise as a new surgical hand scrub formulation with antimicrobial and use characteristics that are significantly improved over current CHG and PVPI formulations. These studies demonstrate the suitability of this formulation for use as a surgical hand scrub and for brushless application.

Comparison of an in vitro skin model to normal human skin for dermatological research.

EpiDerm, an in vitro human skin equivalent (HSE), was compared to normal human breast skin (NHS) to morphologically and biochemically assess its feasibility for dermatological research. Intralot and interlot variability was studied in day 0, 1, 2, and 3 in vitro cultures and in day 0, 3, 5, and 7 NHS. For NHS, light microscopy (LM) at day 0 showed stratified epidermis which exhibited an increase in vacuoles and dark basal cells as storage increased to 3, 5, and 7 days. Transmission electron microscopy (TEM) revealed typical organelles in the epidermis and a convoluted basement membrane at day 0. With increased storage, vacuoles and paranuclear clefts became numerous, necrosis increased, tonofilaments became less organized, and overall cellular integrity decreased. Biochemical data showed consistent MTT and glucose utilization (GU) through day 5, while lactate production decreased to 75% by day 3. By LM, day 0 HSE consisted of a thick, compact, stratum corneum that sent projections between the stratum granulosum cells. By TEM, the configuration organization, differentiation, distribution, and frequency of the organelles differed slightly from NHS. In addition, the basement membrane of the HSE was not completely differentiated, and the dermis was thin and acellular. Although day 1 and 2 cultures showed little change, day 3 exhibited an overall degeneration. Biochemical analysis showed GU and the lactate production decreased through day 3. In conclusion, the EpiDerm HSE, although exhibiting slight differences, was morphologically and biochemically similar to normal human epidermis and may be a valuable model in assessing the toxicology, metabolism, or pharmacology of nonvesicating compounds.

Cutaneous histopathologic features in weanling pigs after exposure to three different doses of liquid sulfur mustard.

Sulfur mustard (2,2' dichlorodiethyl sulfide, HD) is a chemical warfare agent that is easily produced, and may be used against civilian populations as well as against military troops. However, good therapeutic and prophylactic measures await a better understanding of the pathophysiology of lesions produced by this agent. Because the skin remains is one of the principal routes for HD exposure and damage, the study of HD-induced skin lesions is of major interest. Blister formation is a characteristic of HD-induced skin lesions in humans. Attempts have been made to find an animal model that produces cutaneous microblisters after exposure to the naturally occurring liquid as well as vaporized HD. Weanling pigs were exposed to three different doses of liquid HD. Histopathologic findings showed microblister formation as well as variable apoptosis and/or necrosis of epidermal keratinocytes and vascular endothelium. Pig skin is morphologically similar to human skin. In the pig, the epidermal lipids, the density of hair follicles, the presence of sweat glands, the proliferation kinetics, and the antigenicity are all closer to human skin than are rodent models. All these features may be important in lesions induced by HD, and may mean that the pig is a superior model for studying the pathophysiology of HD-induced cutaneous lesions.

Efficacy of tacrine as a nerve agent pretreatment.

Tacrine (THA) was evaluated in vitro and in vivo as a pretreatment for nerve agent intoxication. In vitro experiments showed that the primary effect of THA was direct inhibition of purified fetal bovine serum acetylcholinesterase (AChE) with a slight effect on slowing the aging rate of nerve agent-inhibited AChE. THA produced significant behavioral effects at doses above 1.7 mg/kg, i.m., in the mouse and 3.4 mg/kg, i.m., in the guinea pig. At the no observable effect level (NOEL) for mice (1.7 mg/kg), THA was effective (P < or = 0.05) in reducing tabun- and soman-, but not sarin-induced lethality in mice. Experiments in the guinea pig showed that at the NOEL (3.4 mg/kg, i.m.) THA was not effective in decreasing lethality due to soman exposure. Since there was significant overlap between pharmacologically effective doses of THA and those which produce behavioral toxicity, THA was not considered a suitable pretreatment for nerve agent intoxication.

Acute toxicity, distribution, and metabolism of 2,4,6-trinitrophenol (picric acid) in Fischer 344 rats.

Picric acid (2,4,6-trinitrophenol) is widely used in industry, by the military, and as a research/clinical chemistry reagent. Characterization of the toxicity of this chemical has been limited. Thus the acute toxicity, distribution, and metabolism of picric acid were investigated using Fischer 344 rats. The LD50 for picric acid following oral dosing of male and female rats was established as 290 and 200 mg/kg, respectively. Blood gas analysis indicated severe acidosis during acute intoxication. Metabolism of picric acid was limited to reduction of nitro groups to amines. Metabolites isolated from urine included N-acetylisopicramic acid (14.8%), picramic acid (18.5%), N-acetylpicramic acid (4.7%), and unidentified components (2.4%). Approximately 60% of the parent picric acid was excreted unchanged. The plasma half-life for picric acid was 13.4 h with a gut absorption coefficient (ka) of 0.069 h-1. Twenty-four hours following oral administration of [14C]picric acid (100 mg/kg), the primary depots of radioactivity (per gram tissue basis) were blood, spleen, kidney, liver, lung, and testes. Respective tissue/blood ratios were 0.37, 0.31, 0.28, 0.26, and 0.22. All other tissue assayed had partition ratios < 0.20, with brain and adipose tissue having the least amount of radioactivity. Tissue/blood ratios were essentially maintained over a 48-h postadministration period. Binding (in vitro) of [14C]picric acid to plasma proteins (whole blood preparations) demonstrated both high- and low-affinity binding sites, with dissociation constants of 3.18 x 10(-6) and 2.85 x 10(-4) M, respectively. The findings of this investigation provide information on the acute toxicity, metabolism, and distribution of picric acid, which can be used in risk assessment analyses and in the design of subchronic and chronic toxicity tests.

Evaluation of compounds as barriers to dermal penetration of organophosphates using acetylcholinesterase inhibition.

An efficient, objective method for evaluating the efficacy of barrier compounds in preventing dermal penetration of organophosphates (OP) in rabbits was developed using time-dependent reduction in erythrocyte (RBC) acetylcholinesterase (AChE) activity as an endpoint. Anesthetized rabbits, with or without a dermal application of a mixture of high- and low-molecular-weight polyethylene glycols (mean molecular weight of 540 daltons; PEG 540), were exposed to different percutaneous doses of 3 highly toxic OP compounds. Dose-response curves were generated for RBC AChE inhibition as a function of percutaneous dose for each OP test material over time. From data generated, a single dose of each OP was selected to challenge PEG-540-protected and unprotected animals to validate the method as a means of differentiating effective from ineffective barriers to skin penetration. Data for a complete evaluation of a PEG 540 test barrier application were obtained within 4 h and anesthesia was maintained for the entire period.

Stagewise, adaptive dose allocation for quantal response dose-response studies.

A principal design objective of many dose-response studies is to estimate extreme percentiles of a dose-response distribution, e.g., the ED95 dose for a particular drug therapy, as precisely as feasible using the smallest number of experimental subjects possible. Such a design requirement necessitates that allocation of subjects to drug doses be carried out in a stagewise fashion to maximize the information obtained from each subsequent experimental observation in light of what has previously been determined. This paper describes and illustrates specialized methods and associated computer programs to evaluate, on a stagewise basis, the anticipated relative sensitivities of alternative experimental plans in the case of dichotomous responses. Following each stage of experimentation, the current estimates of the dose-response distribution parameters, as well as the uncertainties in these estimates, are updated and are used to assign subjects to experimental dose levels for the next stage of testing. Competing dose allocations are compared with respect to anticipated improvement in estimation precision. The adoption of such a stagewise dose allocation strategy is illustrated by example.

Stagewise, group sequential experimental designs for quantal responses. one-sample and two-sample comparisons.

The use of stagewise, group sequential experimental designs with dichotomous responses in toxicity or drug screening programs is discussed. Such designs represent a compromise between the standard, fixed sample size designs and fully sequential designs. Stagewise group sequential designs place specified numbers of animals on test at each stage, up to a maximum number of stages. The greatest increases in sample size efficiency occur with small numbers of stages, particularly when going from one stage to two. Two-stage designs can result in a 15 to 20 percent reduction in average sample size. Five-stage designs can result in a 30 to 40 percent reduction in average sample size, with no appreciable decrease in Type 1 error or power. Examples of the efficiencies that arose in actual screening programs are given. This paper demonstrates that the routine use of stagewise, group sequential designs in standardized screening protocols can result in substantial savings in animal use with virtually no sacrifice of statistical sensitivity.

Comparative immunotoxicity of 2,2'-dichlorodiethyl sulfide and cyclophosphamide: evaluation of L1210 tumor cell resistance, cell-mediated immunity, and humoral immunity.

The immunotoxicity of 2,2'-dichlorodiethyl sulfide (sulfur mustard, SM), on humoral and cell-mediated immunity was compared with that of the nitrogen mustard 2-[bis(2-chloroethyl)amino]tetrahydro-2H-1,3,2-oxazophosphorine 2-oxide (cyclophosphamide, CP). SM and CP had similar effects on thymic and splenic weights, spleen cell number, and the formation of antibody producing cells to sheep red blood cells (sRBC) when examined 5 days after exposure, but differed in their effects on body weights. Although there were no differences in the delayed hypersensitivity response to keyhole limpet hemocyanin, CP and SM had different effects in the L1210 tumor cell allograft rejection assay. CP, but not SM, decreased the 28 day survival rate of allogeneic mice exposed to a sublethal L1210 tumor challenge. The differing effects on survival to the L1210 tumor challenge could not be attributed to a direct cytotoxic effect of SM on the L1210 tumor cells as SM did not increase the survival rate or median survival time of syngeneic mice exposed to a lethal L1210 tumor cell challenge. In summary, SM and CP had immunosuppressive effects in the humoral immune assay. Although neither compound suppressed the delayed hypersensitivity response, CP was found to suppress host resistance to L1210 tumor cells.

Metabolism and nephrotoxicity of indan in male Fischer 344 rats.

Indan, a component of fuels, solvents, and varnishes, is metabolized in male Fischer 344 rats to 1-indanol, 2-indanol, 5-indanol, 1-indanone, 2-indanone, 2-hydroxy-1-indanone, cis-1,2-indandiol, and trans-1,2-indandiol. The metabolites were identified using the techniques of gas chromatography (GC) and gas chromatography/mass spectrometry (GC/MS). The rats treated with indan demonstrated the classic lesions of hydrocarbon-induced nephropathy. The kidney damage produced was less than that found for tetralin and other branched-chain acyclic hydrocarbons.

Metabolism and nephrotoxicity of tetralin in male Fischer 344 rats.

Tetralin, a component of fuels, solvents, and varnishes, is metabolized in male Fischer 344 rats to 1-tetralol, 2-tetralol, 2-hydroxyl-1-tetralone, 4-hydroxyl-1-tetralone, 1,2-tetralindiol, and 1,4-tetralindiol. Rats treated with tetralin demonstrated the classic lesions of hydrocarbon-induced nephropathy.

Identification of urinary metabolites in rats exposed to the nephrotoxic agent 2,3,4-trimethylpentane.

Branched-chain hydrocarbon-induced nephropathy in male rats was examined using 2,3,4-trimethylpentane. Lesions are elected only in male rats, not in females or in controls. Mechanisms of nephropathy may be the interaction of metabolites with the male rat-specific protein alpha 2u globulin. The identified urinary metabolites of 2,3,4-trimethylpentane in male rats given the hydrocarbon by gavage are 2,3,4-trimethyl-1-pentanol, 2,3,4-trimethyl-2-pentanol and 2,3,4-trimethyl-1-pentanoic acid. Of the C8-isomers, 2,3,4-trimethylpentane dosing leads to the highest incidence of kidney damage in male rats.

The metabolism of 2,3,4-trimethylpentane in male Fischer-344 rats.

The urinary metabolites of the potent nephrotoxic hydrocarbon 2,3,4-trimethylpentane (2,3,4-TMP) given Fischer-344 male rats by gavage included 1-hydroxy-2,3,4-trimethylpentane, 2,3,4-trimethyl-1-pentanoic acid and 2,3,4-trimethyl-5-hydroxy-1-pentanoic acid. Analyses were performed by gas-liquid chromatography (GC) and gas-liquid chromatography/mass spectrometry (GC/MS). A comparison of the urinary metabolites of 2,3,4-TMP with those of 2,2,4-trimethylpentane (2,2,4-TMP) showed that more monocarboxylic acid was produced with 2,3,4-TMP.

Femoral venipuncture for collection of multiple blood samples in the nonanesthetized rat.

A technique for blood collection via the femoral vein was developed for use in nonanesthetized rats. The technique was useful for single or serial blood collection. Volumes of 0.1 to 1 ml were collected for serum biochemical or venous blood gas determinations. The technique was effective, reproducible, did not require anesthesia, and was less stressful than other methods of blood collection in the rat.

Effect of acetone precipitation on the clinical prediction of respiratory distress syndrome when utilizing amniotic fluid lecithin/sphingomyelin ratios.

Amniotic fluid lecithin-to-sphingomyelin ratios are widely used to predict fetal lung maturity. The inclusion of acetone precipitation in the determination of the lecithin/sphingomyelin ratio has been widely accepted as a necessary step for enhancing predictivity. The lecithin/sphingomyelin ratio of 181 amniotic fluid samples was analyzed with and without acetone precipitation. Sensitivity, specificity, and predictive values were compared. Although there was a significant difference in the numeric value (p less than 0.001) between the two methods, the clinical prediction of the respiratory distress syndrome was not significantly enhanced by acetone precipitation. We conclude that acetone precipitation fails to improve the prediction of fetal lung maturity by lecithin/sphingomyelin ratio analysis.

The metabolism of n-octane in Fischer 344 rats.

The urinary metabolites of n-octane in Fischer 344 rats given the hydrocarbon by gavage included 2-octanol, 3-octanol, 5-oxohexanoic acid, and 6-oxoheptanoic acid. The sex of the animals influenced the relative amounts of metabolites formed. Analyses were performed by gas-liquid chromatography (GC) and gas-liquid chromatography/mass spectrometry (GC/MS). This is the first reported finding of keto acids in hydrocarbon oxidative metabolism. No kidney damage was found as a result of n-octane dosing although the 2,2,4-trimethylpentane (iso-octane) isomer does cause kidney lesions in male rats.

A subchronic dermal exposure study of diethylene glycol monomethyl ether and ethylene glycol monomethyl ether in the male guinea pig.

Diethylene glycol monomethyl ether (DEGME) has been selected as a replacement anti-icing additive for ethylene glycol monomethyl ether (EGME) in Navy jet aircraft fuel. This experiment was performed to determine whether DEGME produced similar toxicity to EGME following dermal exposure. Male guinea pigs were dermally exposed to 1.00, 0.20, 0.04, or 0 (control) g/kg/day DEGME for 13 weeks, 5 days/week, 6 hr/day. Another group of animals was similarly exposed to 1.00 g/kg/day EGME. Body weights as well as testicular and splenic weights were reduced as a result of exposure to EGME, DEGME-exposed animals exhibited decreased splenic weight in the high- and medium-dose (1.00 and 0.20 g/kg/day) exposure groups only. Hematologic changes in EGME-exposed animals included mild anemia with increased erythrocytic mean corpuscular volumes and a lymphopenia with increased neutrophils. Similar hematological changes were not observed in any animals exposed to DEGME. Serum creatine kinase activity was increased in animals exposed to EGME, and serum lactate dehydrogenase activity was increased in EGME and 1.00 g/kg/day DEGME-exposed animals. In general, DEGME produced minimal toxicological changes following dermal exposure, whereas the toxicological changes observed following similar exposure to EGME were much more profound.

Identification of urinary metabolites of the nephrotoxic hydrocarbon 2,2,4-trimethylpentane in male rats.

The compound 2,2,4-trimethylpentane (2,2,4 TMP) is reported to be especially potent in inducing kidney lesions in male rats (1,2). Although the pathology produced by 2,2,4 TMP has been examined (1), there are no reports concerning the metabolism of 2,2,4 TMP by the male rat. The eight principal urinary metabolites of 2,2,4 TMP found in the urine of Fischer 344 male rats are: 2,2,4-trimethyl-1-pentanol, 2,4,4-trimethyl-1-pentanol, 2,4,4-trimethyl-2-pentanol, 2,2,4-trimethyl-1-pentanoic acid, 2,4,4-trimethyl-1-pentanoic acid, 2,4,4-trimethyl-2-hydroxy-1-pentanoic acid, 2,2,4-trimethyl-5-hydroxy-1-pentanoic acid and 2,4,4-trimethyl-5-hydroxy-1-pentanoic acid.