Activities of hepatic cytosol enzymes involved in UDP-g1ucuronic acid synthesis as well as in glutathione reduction and conjugation systems were determined following administrations of butylated hydroxyanisole (approximately 5 mmol/kg body weight/day) and of equimolar intake doses of its structural anglogs. These compounds included the multi-functional group side chain compounds (t-butyl hydroquinone, 4-hydroxy- anisole, hydroquinone, benzoquinone) and the mono-functional side chain compounds (t-butyl benzene, anisole, phenol). They were administered to mice for 10 days either by mixing them in the diet or by oral intubations. Results showed that glutathione Stransferase activities were markedly increased by all tested compounds except for the t-butyl benzene. Activities of glutathione reductase and glucose 6-phosphate dehydrogenase were increased together on1y by BHA and t-butyl hydroguinone. UDP-glucose dehydrogenase and NADH:quinone reductase activities were significantly elevated by the multi-functional side chain compounds, but not by the mono-functional analogs. The relations between chemical structures of tested BHA analogs and elevations of the measured hepatic cytosol conjugation (detoxification) system enzyme activities for the metabolism and excretion of BHA analogs are discussed.
Animal ; Anisoles/metabolism* ; Butylated Hydroxyanisole/analogs & derivatives ; Butylated Hydroxyanisole/metabolism* ; Cytosol/enzymology* ; Glutathione/metabolism* ; Mice ; Uridine Diphosphate Glucuronic Acid/biosynthesis* ; Uridine Diphosphate Sugars/biosynthesis*

OBJECTIVEIn order to explore the pathway of dealkylation of pesticides other than cytochrome P450 monocoxygenases, lipoxygenase (LOX)-mediated demethylation of aminocarb and some other pesticides were measured.

METHODFormaldehyde generated in the reaction was estimated by Nash reaction to express the rate of demethylation of pesticides mediated by soy lipoxygenase (SLO).

RESULTSN-demethylation of aminocarb mediated by SLO was found to depend on the incubation time, concentration of the enzyme, concentration of aminocarb and hydrogen peroxide. Under optimal conditions, Vmax value of 18 nmol of formaldehyde.min-1.nmol-1 of lipoxygenase was observed. The reaction exhibited Km values of 3.4 mmol/L for aminocarb and 235 mumol/L for hydrogen peroxide. A strong inhibition of the reaction by nordihydroguaiaretic acid, gossypol, and phenidone clearly implicated the lipoxygenase involvement as the protein catalyst. A significant decline in the formaldehyde accumulation in the presence of either reduced glutathione or dithiothreitol suggested generation of a free radical species as an initial oxidation intermediate during the demethylation of aminocarb by SLO. The inhibition of formaldehyde generation by butylated hydroxyanisole(BHT) and butylated hydroxy toluene(BHA) further supported this contention. In addition to aminocarb, seven other pesticides were also found to undergo N-demethylation, albeit at relatively low rates.

CONCLUSIONCertain pesticides may oxidatively undergo dealkylation via the lipoxygenase pathway in animals and plants.

Butylated Hydroxyanisole ; pharmacology ; Butylated Hydroxytoluene ; pharmacology ; Dealkylation ; Free Radicals ; Lipoxygenase ; physiology ; Pesticides ; metabolism ; Phenylcarbamates ; metabolism ; Soybeans ; enzymology

Effects of feeding 2(3)-tert-butyl 4-hydroxyanisole (BHA) and 3, 5-di-tert-butyl 4-hydroxytoluene (BHT) on the rates of mixed function oxidation and conjugation enzyme reactions have been determined using isolated hepatic microsomal fractions and isolated perfused livers of mice. The treatments with either of the antioxidants have increased the rates of O-demethylation for p-nitroanisole and of O-deethylation for 7-ethoxycoumarin up to 2-fold, both in microsomes and in perfused liver. Analysis of the perfusate showed that the increased amounts of p-nitrophenol and 7-hydroxycoumarin produced by the elevated mixed-function oxidase activities were reflected by the increase in the amounts of glucuronide conjugates and not in the increase for the amounts of the sulfate ester conjugates. Comparison of results also indicated that in the perfused liver, the maximal rate of metabolite conjugation is limited by the maximal rates of the initial mixed function oxidase activities.
Alkylation ; Animal ; Anisoles/metabolism ; Anisoles/pharmacology* ; Butylated Hydroxyanisole/administration & dosage ; Butylated Hydroxyanisole/pharmacology* ; Butylated Hydroxytoluene/administration & dosage ; Butylated Hydroxytoluene/analogs & derivatives* ; Butylated Hydroxytoluene/pharmacology ; Comparative Study ; Coumarins/metabolism ; Female ; Glucuronosyltransferase/metabolism ; Liver/metabolism* ; Mice ; Microsomes, Liver/enzymology ; Microsomes, Liver/metabolism* ; Mixed Function Oxygenases/metabolism ; Oxidation-Reduction ; Perfusion ; Support, U.S. Gov't, P.H.S.

AIMTo study the mechanism of butylated hydroxyanisole-induced neural differentiation of fetal liver cells in vitro.

METHODS14.5-day-old mouse fetal liver-derived cells were cultured, and were induced by 200 micromol/L butylated hydroxyanisole (BHA) combined with PI3K inhibitor LY294002 (20 micromol/L), and then were incubated in serum-free medium. Expression of genes in treated or untreated cells were assayed by Western blotting or RT-PCR.

RESULTSThere was low level of neurofilament-L (NF-L) and brain factor-1 (BF-1) but no neurofilament-H (NF-H) and tyrosine hydroxylase (TH) in fetal liver cells. BHA promoted significantly expression of neuron-specific NF-L, NF-H, BF-1, and TH in fetal liver cells. NF-L mRNA increased 5.8 fold, NF-H mRNA 8.0 fold, BF-1 mRNA 2.68 fold, and TH mRNA 30 fold, respectively (all P < 0.01 vs untreated cells). NF-L protein increased 11.29 fold, NF-H 5.5 fold, BF-1 2.53 fold, TH 4.76 fold. Moreover, expression of these BHA-induced genes were inhibited by PI3K inhibitor LY294002.

CONCLUSIONBHA induced neural differentiation of fetal liver cells through PI3K.

Animals ; Butylated Hydroxyanisole ; pharmacology ; Cells, Cultured ; Embryo, Mammalian ; cytology ; Hepatocytes ; drug effects ; metabolism ; Mice ; Mice, Inbred C57BL ; Nerve Tissue Proteins ; biosynthesis ; genetics ; Phosphatidylinositol 3-Kinases ; metabolism

An experiment was conducted in order to investigate the effect of p-dimethylaminoazobenzene (DAB) and 2(3)-tert-butyl-4-hydroxyanisole (BHA) on the lipid peroxidation and peroxide-destroying enzyme system in the rat liver. Dietary supplementation of DAB (0.06%) for three weeks caused the elevation of glutathione-S-transferase activity by 60% and glutathione reductase by 50%, but it decreased glutathione peroxidase and catalase activities significantly. Dietary supplementation of BHA (0.75%) also increased glutatione-S-transferase activity in the liver by 2 folds, and it counteracts DAB effect on the glutathione peroxidase and catalase activities. There was a marked increase in malon-dialdehyde content in the postnuclear fraction of liver by the treatment of DAB, but the addition of BHA lowered the malondialdehyde content to almost the control level. The protective effect of BHA on the lipid peroxidation induced by DAB administration at the enzyme level seems to be due to the induction of glutathione-S-transferase and the protection of glutathione peroxidase and catalase activities from being lowered by DAB administration.
Animal ; Anisoles/pharmacology* ; Butylated Hydroxyanisole/pharmacology* ; Glutathione Peroxidase/analysis* ; Glutathione Reductase/analysis* ; Glutathione Transferase/analysis* ; Lipid Peroxides/metabolism* ; Liver/drug effects* ; Liver/metabolism ; Male ; Peroxidases/analysis* ; Rats ; p-Dimethylaminoazobenzene/pharmacology*