Floyd R. Fullerton

Influence of oral administration of sulfamethazine on thyroid hormone levels in fischer 344 rats

Fischer 344 rats (810 of each sex) were divided into treatment groups and fed diets containing 0, 10, 40, 600, 1200, or 2400 ppm sulfamethazine. Serum samples were analyzed for levels of thyroid-stimulating hormone (TSH), total thyroxine (T4), total triiodothyronine (T3), and T3 uptake after 12, 18, or 24 mo of continuous dosing. There were no statistically significant differences in T3 levels or percent T3 uptake for either sex after any of the exposure periods. The serum T4 levels were lower (p less than 0.05) for females dosed at 1200 and 2400 ppm for 18 mo and for males dosed at 600, 1200, or 2400 ppm sulfamethazine for 24 mo than for those dosed at levels of 40 ppm or less. Serum TSH levels showed a general increasing trend (but not statistically significant) among animals receiving 600 ppm or more sulfamethazine. There was a significant dose-related reduction in (T3 + T4)/TSH ratio for both sexes (p less than 0.05) after 18 and 24 mo of exposure at dose levels of 600 ppm or more. A lack of response at 12 mo may have been due to the shorter treatment time. At each sacrifice period both sexes of rats fed sulfamethazine at 1200 and 2400 ppm had significantly heavier (p less than 0.05) thyroid weights than animals fed control diet. The heavier thyroid weights in the dosed animals may have resulted from increased TSH levels. The cause of reduction in serum T4 was not clearly evident. Therefore, the thyroid hormone to pituitary feedback mechanism apparently compensated for sulfamethazine effects in most animals. This would suggest that the thyroid gland was not irreversibly affected.

Analytical procedure for determination of S-adenosylmethionine, S-adenosylhomocysteine, and S-adenosylethionine in same isocratic HPLC run, with a procedure for preparation and analysis of the analog S-adenosylhomocysteine sulfoxide

Abstract Virtually all methyltransferase enzymes are regulated largely by the relative levels of S-adenosylmethionine (SAM) to its metabolic product, S-adenosylhomocysteine (SAH). Ethionine is the hepatocarcinogenic antimetabolite of methionine, and has been found to produce hypomethylation of hepatic DNA when fed to rats in acute doses. The hypomethylation apparently results from the accumulation in the liver of S-adenosylethionine (SAE), the sulfur activation product of ethionine, which is a competitive inhibitor of DNA methylase. Researchers seeking to measure SAM and SAH levels by HPLC in the past have experienced numerous analytical problems because of their separation characteristics. Previous methods have either required two separate HPLC runs or used gradient elution to measure the two compounds. The method outlined here, is an accurate and precise method, that measures SAM and SAH as well as SAE in a single isocratic HPLC run. S-Adenosyl-l-homocysteine sulfoxide (SAHO), the sulfoxide of SAH is kn...

Increased incidence of spontaneous and 2-acetylaminofluorene-induced liver and bladder tumors in B6C3F1 mice fed AIN-76A diet versus NIH-07 diet

Diet is a major influence on the responses of experimental animals to drugs, toxins, and carcinogens. Two diets used widely in toxicological and carcinogenic studies, and considered to be nutritionally adequate, were compared with respect to neoplastic responses to 2-acetylaminofluorene (2-AAF). Both sexes of weanling B6C3F1 mice were fed either AIN-76A (a purified diet) or NIH-07 (a natural ingredient diet), with or without 2-AAF, for up to 2 years. Dosages of 2-AAF were administered to males at 0, 40, 60, or 80 ppm in each diet and to females at 0, 150, 200, or 250 ppm. Each group consisted of 96 mice, except the groups of females dosed at 0 and 150 ppm, which consisted of 120 and 72 mice, respectively. The incidence of malignant liver tumors was significantly greater in all AIN-fed groups compared to corresponding NIH-fed groups, as was the total incidence of tumors (malignant + benign). Similarly, the incidence of malignant and total bladder tumors was greater in AIN-fed groups of mice administered the two high doses of 2-AAF for each sex compared to NIH-fed groups. No malignant bladder tumors were observed among any of the groups of mice receiving control diets. This was also true for the males on low dosages (40 ppm) of 2-AAF. The AIN-fed group of low dose females (150 ppm) developed 4% incidence of malignant bladder tumors. This study dramatically showed the importance of diet selection in chronic carcinogenic studies and suggests that results obtained with the use of a purified diet may differ both qualitatively and quantitatively from results obtained with a natural ingredient diet.

Effects of Diet Type on Incidence of Spontaneous and 2-Acetylaminofluorene-Induced Liver and Bladder Tumors in BALB/c Mice Fed AIN-76A Diet versus NIH-07 Diet

Diet is a major influence on the responses of experimental animals to drugs, toxins, and carcinogens. Two diets used widely in toxicological and/or nutritional studies, and considered to be nutritionally adequate, were compared with respect to their influence on growth, body weight, lifespan, spontaneous neoplasia, and neoplastic responses to 2-acetylaminofluorene (2-AAF). Both sexes of weanling BALB/c mice were fed either a purified diet (AIN-76A) or a nonpurified, natural ingredient diet (NIH-07), with or without 2-AAF for up to 2 years. Dosages of 2-AAF were administered to males at 0, 20, 40, or 60 ppm in each diet and to females at 0, 100, 125, or 150 ppm. Each group consisted of 96 mice. In most instances, males and females fed purified diet (AIN-fed) gained weight more rapidly, attained higher maximum body weights, and died earlier than their non-purified diet (NIH-fed) counterparts. 2-AAF inhibited weight gain significantly only in AIN-fed females. Thus, females receiving 150 ppm 2-AAF gained little more than their NIH-fed counterparts. At the dosages used in males, 2-AAF did not induce liver neoplasia but the AIN diet was clearly associated with a higher spontaneous frequency of liver neoplasia than the NIH diet. Although 2-AAF induced liver tumors in females fed either diet at all dosages, a higher frequency and earlier appearance of liver tumors among AIN-fed females than their NIH-fed counterparts was apparent mainly at the lowest dosage. 2-AAF induced bladder neoplasia in both sexes.(ABSTRACT TRUNCATED AT 250 WORDS)

Determination of 2-acetylaminofluorene and its metabolites in urine by high pressure liquid chromatography.

Abstract 2-Acetylaminofluorene, a well-documented carcinogen, is used extensively as a positive control in the bioassay of numerous suspect carcinogens. Consequently, there is a need for a rapid, sensitive analytical method for determining 2-acetylaminofluorene and its metabolites in urine. Such a method would be useful in monitoring workers for potential exposure as well as for determining the applicability of the test species by metabolism studies. A high pressure liquid chromatography method for the identification and quantitation of 2-acetylaminofluorene and its major metabolites in urine has been developed. The primary obstacle in developing the method was the presence of large amounts of endogenous urinary components in ether extracts of urine having chromatographic properties similar to 2-acetylaminofluorene and its metabolites. This problem was obviated by the use of a combination of Carbowax 400 and C18/Corasil columns. N-Hydroxy-2-acetylaminofluorene was determined directly from the Carbowax 400 column. Fractions containing 2-acetylaminofluorene and its 1-, 3-, 5-, and 7-hydroxy metabolites were then separated from interfering materials and quantitated by fractionation on a C18/Corasil column. The response of the system was linear down to 100 ng of N-hydroxy-2-acetylaminofluorene and to 10 ng of 2-acetylaminofluorene and the other metabolites.

Comparison of histological responses of balb/C and B6C3F1 female mice to estradiol when fed purified or natural‐ingredient diets

Female BALB/c and B6C3F1 mice were examined after a 3-wk exposure to dietary estradiol (0, 400, 800, 1600, and 3200 ppb) in a purified diet (AIN-76A) or a natural-ingredient diet (NIH-07). Histological findings, which became more prevalent with increasing estradiol dosage in both mouse genotypes, included vaginal hyperkeratosis and mucoid stroma, uterine inflammation, hydrometra and glandular hyperplasia, and ovarian corpora lutea depletion. At the two lower doses of estradiol, responses were generally more prevalent or severe in mice fed the purified diet than in those fed the natural-ingredient diet. However, in BALB/c mice, several responses to the two higher estradiol doses were greater when estradiol was given in the natural-ingredient diet rather than in the purified diet. These responses included corpora lutea depletion, vaginal hyperkeratosis, and uterine inflammation and hydrometra. In B6C3F1 mice, most responses to estradiol at concentrations of 400, 800, and 1600 ppm were more prevalent or severe in mice fed the purified diet than in those fed the natural-ingredient diet. It can be concluded that several responses to estradiol in mice maintained for a 3-wk period on a purified diet differed significantly from mice maintained on a natural-ingredient diet.

Effects of purified (AIN‐76A) and natural‐ingredient (NIH‐07) diets on responses of BALB/c and B6C3F1 female mice to estradiol

Female BALB/c and B6C3F1 mice were examined after a 3-wk exposure to dietary estradiol (0, 400, 800, 1600, and 3200 ppb) in a purified (AIN-76A) or a natural-ingredient (NIH-07) diet. The use of AIN-76A was associated with a 9-13% greater (p less than 0.001) body weight and a 36-43% higher (p less than 0.001) serum cholesterol in both mouse genotypes when compared to mice fed NIH-07. Conversely, when fed NIH-07, both mouse genotypes had a 20-22% higher (p less than 0.003) serum urea nitogren and 2-3.5% higher erythrocyte count (p less than 0.001) and hemoglobin concentration (p less than 0.04) than when fed AIN-76A. Reduced erythrocyte parameters suggest that chronic feeding of the purified diet might result in anemia. No significant compound or diet-related differences were noted for serum creatinine, alanine aminotransferase, aspartate aminotransferase, or gamma-glutamyl transferase. Although there was no diet effect on absolute or differential white blood cell count, estradiol caused a decrease in the total white blood cell count (p less than 0.014) and an increase in the percentage of polymorphonuclear leukocytes (p less than 0.014) in BALB/c and decreased the percentage of lymphocytes (p less than 0.005) in B6C3F1 females. In addition, estradiol increased uterine weight and inhibited thymic and splenic weights in one or both genotypes. Spleen and thymus weight responses to estradiol were not significantly influenced by diet. However, the uterine weight responses to estradiol were apparently influenced by diet in both genotypes. In B6C3F1 mice, the uterus weighed more at each level of estradiol when mice were fed AIN-76A compared to NIH-07 diet. In BALB/c mice, this was true only at the two lower dietary concentrations of estradiol. In conclusion, mice fed the purified diet, AIN-76A, differed from those fed the cereal-based diet, NIH-07, in hematology, clinical chemistry, and uterine weight response to estradiol.

Effect of nitrous oxide on exposure on maternal and embryonic S-adenosylmetthionine levels and ornithine decarboxylase activity

Nitrous oxide is suspected to be a developmental toxicant in humans. The anesthetic does produce increases in the resorption and malformation frequencies in rodents. The mechanism for the drugs developmental toxicant effects is unknown. Embryonic DNA synthesis is decreased; however, this decrease does not appear to be due to depressed levels of adenine or guanine. In this investigation, we examined the effect of N2O on maternal and embryonic S-adenosylmethionine (AdoMet) levels and ornithine decarboxylase (ODC) activity, and the effect of exogenous methionine (Met) on these parameters was also examined. AdoMet and ODC are involved in polyamine synthesis, and polyamines are involved in regulation of macromolecular synthesis. Pregnant rats were treated with N2O for 24 hours beginning on the morning of day 10 of gestation. There was no effect of N2O on maternal hepatic AdoMet or S-adenosylhomocysteine (AdoHcy) levels; there was also no effect on embryonic AdoMet. Embryonic AdoHcy could not be detected in many of the samples; however, N2O treatment did significantly increase the number of embryonic samples in which AdoHcy was detectable. ODC activity was not affected by either treatment in dams but was increased by N2O in embryos. It is possible that the embryotoxic effect of this anesthetic is mediated by alterations in the AdoMet to AdoHcy ratio or to changes in ODC activity and polyamine synthesis.