Effect of the flavonoid baicalein as a feed additive on the growth performance, immunity, and antioxidant capacity of broiler chickens.

Baicalein, the main flavonoid extracted from the root of Scutellaria baicalensis Georgi, has been demonstrated to exert multiple pharmacological effects, and thus could be utilized as a potential feed additive in broiler chickens. This study evaluated the effects of broiler chicken diet supplementation with baicalein on growth performance, immunity, and antioxidant activity at levels of 100 and 200 mg/kg. No significant effect on average daily feed intake (P > 0.05) of broilers with diets supplemented with baicalein was found compared to those on the basal diet or butylated hydroxytoluene (BHT) during the 35-d feeding trial. The addition of baicalein to the basal diet significantly increased average body weight, body weight gain, average weight gain, and the feed conversion ratio of birds during 21 to 42 d and 7 to 42 d of age, respectively. The best numerical values for the overall growth performance were observed in broilers fed on diets containing 200 mg/kg of baicalein. Baicalein supplementation significantly increased the ratio of CD3+/CD4+ and CD3±/CD8+, the concentration of IFN-γ, anti-IB antibody titer, and the spleen index compared with the control group (P < 0.05). Total cholesterol, the ratio of non-HDL-C/HDL-C, LDL-C/HDL-C, TC/HDL-C, triglycerides, and low-density lipoprotein cholesterol were significantly decreased after intake of baicalein compared with both the basal diet and the BHT-supplemented diet, whereas the SOD, GSH-Px, and CAT activity in the serum increased with the supplementation of baicalein. The T-AOC activity, T-SOD, and GSH-Px level in liver tissues was significantly increased by inclusion of baicalein, and intake of baicalein or BHT significantly decreased the malondialdehyde level found both in serum and meat tissue. Thus, the results obtained here indicate that the baicalein can be used as an effective natural feed additive in broiler chicken diets, and that 100 to 200 mg/kg can be considered as the optimum dosage.

Effect of butylated hydroxyl toluene on the immune response of Rift Valley fever vaccine in a murine model.

The present study was planned to examine the effect of butylated hydroxy toluene (BHT) on the immune response of Rift Valley fever vaccine (RVFV) in Swiss mice. Animals were divided into four equal groups. The first group was kept as negative control. The 2nd group was orally administrated with the acceptable daily intake (ADI) of BHT 0.3 mg/kg b.wt. daily for 21 days and the 3rd group were vaccinated only by inactivated RVFV at a dose of 0.2 ml I/P two times. The 4th group was orally administrated BHT as in the 2nd group and vaccinated by inactivated RVFV as in the 4th group. Blood samples were collected from all groups two weeks from booster vaccination. The cellular immunity was determined by leucocytic indices and the neutrophil-lymphocyte ratio (NLR) whereas, humoral immunity was evaluated with IgG antibodies titer using enzyme-linked immune-sorbent assay (ELISA) test, serum neutralization test (SNT) and challenge test. BHT induced leucopenia, neutrophilia and marked lymphocytopenia in both non-vaccinated and vaccinated mice. Moreover, BHT significantly decreased the efficiency of vaccination by inducing 70% cytopathic effect (CPE) in the infected cell cultures and increasing the ED50 value of RVFV vaccine. The present study indicates that BHT possesses a potential for decreasing both cellular and humoral mediated mechanisms.

Dietary butylated hydroxytoluene improves lipid metabolism, antioxidant and anti-apoptotic response of largemouth bass (Micropterus salmoides).

A 10-week growth trail was conducted to investigate the efficacy and tolerance of dietary butylated hydroxytoluene (BHT) by evaluating inflammation, apoptosis and hepatic disease related to oxidative stress in largemouth bass (Micropterus salmoides). Four experimental diets were prepared with BHT supplement levels of 0 (B0), 150 (B150), 300 (B300) and 1500 (B1500) mg/kg, in which B150 was at the maximum recommended level established by European Union Regulation, and the B300 and B1500 levels were 2 and 10-fold of B150, respectively. Each diet was fed to 6 replicates with 30 largemouth bass (initial body weight, IBW = 6.20 ± 0.01 g) in each tank. The BHT inclusion level did not affect the specific growth rate, but fish in the B150 group showed the lowest feed conversion rate (P < 0.05). BHT inclusion significantly decreased the levels of plasma TC, TG, LDL, ALT and AKP, and increased the (HDL-C)/TC ratio (P < 0.05). Plasma MDA was significantly decreased in the B150 group and GSH-Px was extremely enhanced in each BHT inclusion group (P < 0.05). Hepatic T-AOC was significantly enhanced and O2- was significantly decreased in each BHT inclusion group compared to the B0 group (P < 0.05), as well as hepatic MDA was significantly decreased in B1500 group (P < 0.05). Dietary BHT inclusion down-regulated the hepatic mRNA levels of inflammation, apoptosis and fibrosis related genes, including TNFα, TGF-ß1, α-SMA, IL8, IL11ß and caspase-9. Moreover, BHT could improve hepatic lipid metabolism via up-regulating the mRNA levels of APOA1, CYP7A1, CYP8B1, and down-regulating the mRNA levels of PPAR-γ and APOB. Histological examination of the liver morphology with H&E and Sirius Red staining showed that BHT inclusion decreased necrotic degenerative changes and collagen deposition in largemouth bass. An immunofluorescence examination revealed significantly decreased cleaved caspase-3 signals in the BHT groups. In conclusion, the results demonstrated that ROS induces hepatic cell apoptosis and fibrosis via the intrinsic pathway of apoptosis by activating caspase-9 in the mitochondria and then initiates apoptosis by activating caspase-3. Consuming 2.32-23.80 mg/kg·bw/d (150-1500 mg/kg in diet) of BHT effectively improved the plasma and hepatic lipid metabolism, antioxidant response as well as reduced ROS production, protecting hepatic cells from injury. It is implied that even a 10-fold increase of the maximum level of BHT (150 mg/kg) is safe for the largemouth bass.

Activation of liver X receptor inhibits the development of pulmonary carcinomas induced by 3-methylcholanthrene and butylated hydroxytoluene in BALB/c mice.

We previously reported that LXR ligand, T0901317, inhibited the growth of inoculated Lewis lung carcinoma in C57BL/6 mice by activating IFN-γ production. However, the effects of T0901317 on carcinogen-induced pulmonary carcinomas remain unknown. In this study, we initially conducted a statistical analysis on the data of human lung cancer samples extracted from the TCGA database, and determined that survival rate/time of lung cancer patients and grade of lung adenocarcinoma were positively and negatively related to lung IFN-γ levels, respectively. We then determined the inhibitory effects of T0901317 on mouse pulmonary carcinomas induced by 3-methylcholanthrene (MCA) and butylated hydroxytoluene (BHT) or urethane. We found that T0901317 reduced morbidity and mortality in MCA/BHT-injected BALB/c mice by inhibiting lung adenocarcinoma. T0901317 also protected C57BL/6 mice, but not IFN-γ deficient (IFN-γ(-/-), C57BL/6 background) mice, against MCA/BHT-induced lung hyperplasia/inflammation. In addition, we determined that T0901317 inhibited urethane-induced lung tumors in BABL/c mice. Furthermore, we determined that T0901317 prevented metastasis of 4T1 breast cancer cells in BALB/c mice. Administration of T0901317 substantially increased serum IFN-γ levels and lung IFN-γ expression in BABL/c and C57BL/6 mice. Taken together, our study demonstrates that LXR inhibits MCA/BHT-induced pulmonary carcinomas in BABL/c mice and the inhibition is associated with induction of IFN-γ production.

Influence of BHT inclusion on post-thaw attributes of human semen.

The aim of this study was to determine the effect of butylated hydroxytoluene (BHT) supplemented cryopreservation medium on sperm parameters during the freeze-thaw process. In addition, sperm lipid peroxidation, DNA damage, and the amount of reactive oxygen species (ROS) were determined. Semen samples were obtained from 75 donors. Fifteen semen samples were used for optimizing BHT concentration and incubation time and 60 samples were used for the final experiments. After the determination of basic parameters, groups of three sample with similar parameters were pooled and processed by Pure Sperm gradient centrifugation. The semen samples were then diluted with normal freezing medium (control) or a medium containing 0.5 mM BHT (test) for 5 minute and stored in liquid nitrogen. Frozen cryovials were thawed individually for 20 seconds in a water bath (37°C) for evaluation. Freezing extenders supplemented with 0.5 mM BHT led to higher sperm motility and viability compared with control samples (p < 0.001). Furthermore, the addition of BHT decreased malondialdehyde (MDA) formation, DNA fragmentation, and ROS content compared with controls (p < 0.001). Our results showed that the addition of BHT to the freezing medium could be of advantage in reducing ROS and preventing the detrimental effect of ROS on the human sperm function.

Effect of antioxidant mixtures on growth and ochratoxin a production of Aspergillus section Nigri species under different water activity conditions on peanut meal extract agar.

The effect of mixtures of antioxidants butylated hydroxyanisol (BHA) and propyl paraben (PP) on lag phase, growth rate and ochratoxin A (OTA) production by four Aspergillus section Nigri strains was evaluated on peanut meal extract agar (PMEA) under different water activities (a(w)). The antioxidant mixtures used were: BHA + PP (mM), M1 (0.5 + 0.5), M2 (1.0 + 0.5), M3 (2.5 + 0.5), M4 (0.5 + 1.0), M5 (1.0 + 1.0), M6 (2.5 + 1.0), M7 (5.0 + 2.5) and M8 (10 + 2.5). The mixture M8 completely suppressed mycelial growth for all strains. A significant stimulation in OTA production was observed with mixtures M1 to M5 mainly at the highest a(w); whereas M6, M7 and M8 completely inhibited OTA production in all strains assayed; except M6 in A. carbonarius strain (RCP G). These results could enable a future intervention strategy to minimize OTA contamination.

Effects of salt, BHA/BHT, and differing phosphate types on quality and sensory characteristics of beef longissimus muscles.

USDA Select striploins (n = 20) were cut into thirds (anterior, medial, and posterior) and randomly assigned to 1 of 6 treatments. Treatments included: (1) control (C); (2) 0.006% BHA (butylated hydroxyl anisole)/BHT (butylated hydroxytoluene) (70%/30%) (BB); (3) 0.4% trisodiumphosphate (CT); (4) 0.4% sodiumtripolyphosphate with 0.5% salt (BH); (5) sodiumtripolyphosphate, 0.5% salt, and 0.006% BHA/BHT (70%/30%) (SB); (6) 0.2% sodiumtripolyphosphate, 0.2% trisodiumphosphate, and 0.5% salt (STB). Muscle sections were injected to 110% (10% pump) of their weight with their respective treatments. Inclusion of BHA/BHT allowed for lower mean oxidation values. Regardless of phosphate type, muscles treated with both phosphate and salt had lower retail purge (P < 0.05). Sensory panelists rated (P < 0.05) STB, SB, and BH to be juicier than all other treatments. These data suggest that inclusion of both salt and phosphate can enhance palatability, lower cook loss, and retail purge.

Butylated hydroxytoluene inclusion in semen extender improves the post-thawed semen quality of Nili-Ravi buffalo (Bubalus bubalis).

The study was carried out to evaluate the potential impact of butylated hydroxytoluene (BHT) on the frozen-thawed semen quality of Nili-Ravi buffalo bulls. Ejaculated bull semen was extended in a Tris-citrate egg yolk extender containing various concentrations of BHT (0.5, 1.0, 2.0 and 3.0mM). Semen was frozen at -196 degrees C using 50 x 10(6) spermatozoa per 0.5 mL straws. Five straws from each treatment were thawed to assess the semen quality in terms of sperm motility, viability, plasma membrane integrity and acrosomal integrity. Post-thawed sperm motility was determined using a phase-contrast microscope. Viability, plasma membrane integrity and acrosomal integrity were evaluated by the supravital staining, hypo-osmotic swelling test and normal acrosomal reaction, respectively. The highest (P<0.05) motility, acrosomal integrity and hypo-osmotic swelling response of spermatozoa was achieved by addition of 1.0 and 2.0mM BHT to semen extender. However, highest (P<0.05) viability of spermatozoa was achieved by inclusion of 2.0mM BHT. The higher concentration of BHT (3.0mM) reduced the motility, acrosomal integrity, viability and hypo-osmotic swelling response of the spermatozoa compared to other concentration used. In conclusion, BHT when added in the semen extender can improve the semen quality of buffalo bulls.

Mechanisms of butylated hydroxytoluene chemoprevention of aflatoxicosis--inhibition of aflatoxin B1 metabolism.

Chemoprevention of toxicoses and/or cancer through the use of nutrients or pharmacologic compounds is the subject of intense study. Among the many compounds examined, food additives such as antioxidants are being considered due to their ability to reduce disease formation by either induction or inhibition of key enzyme systems. One such compound, butylated hydroxytoluene (BHT), has been found to protect against cancer formation caused by exposure to aflatoxin B1 (AFB1) in rodents. We have shown that dietary BHT protects against clinical signs of aflatoxicosis in turkeys, a species that is very susceptible to this mycotoxin. In this study, the effect of BHT on AFB1 metabolism and other cytochrome P450 (CYP)-related enzyme activities in turkey liver microsomes was examined to discern possible mechanisms of BHT-mediated protection against aflatoxicosis. Ethoxyresorufin O-deethylase (EROD), methoxyresorufin O-demethylase (MROD), prototype activities for CYP1A1 and 1A2, respectively, were decreased in the BHT fed (4000 ppm) animals, while oxidation of nifedipine, a prototype activity for CYP3A4, was increased. However, BHT added to microsomal incubations inhibited these CYP activities in a concentration-related manner. Importantly, BHT inhibited conversion of AFB1 to the reactive intermediate AFB1-8,-9-epoxide (AFBO), exhibiting Michaelis-Menton competitive inhibition kinetics (Ki=0.81 microM). Likewise, microsomes prepared from turkeys fed BHT were significantly less active in AFBO formation compared to those from control birds. When turkeys were fed BHT for up to 40 days, residual BHT was present in liver, breast meat, thigh meat and abdominal fat in concentrations substantially below U.S. FDA guidelines for this antioxidant, but in concentrations greater than the Ki, likely sufficient to inhibit bioactivation of AFB1in vivo. BHT-induced hydropic degeneration in the livers of BHT fed animals was significantly greater in birds that remained on BHT treatment for up to 30 days, but this lesion diminished in animals fed for 40 days or when returned to a control diet. The data indicate that the observed chemopreventive properties of BHT in turkeys may be due, at least in part, to its ability to inhibit hepatic AFB1 epoxidation and also that the BHT-induced hydropic degeneration is reversible and does not appear to cause long-term effects.

Role of maternal dietary antioxidant supplementation in murine placental and fetal limb development.

Methylnitrosourea (MNU) is a multisystem teratogen that damages proliferating cells through macromolecule alkylation and generation of reactive oxygen species (ROS). Murine dams exposed to MNU midgestation produce offspring with distal limb malformations, an outcome reduced by maternal immune stimulation. Immunostimulatory effects of antioxidant therapy may in part explain this improved birth outcome. The present study hypothesizes that placental, rather than fetal, damage from excessive ROS may contribute to MNU-induced embryopathy. Fetal limbs and placentas were examined in immunotolerant CD-1 and immunosensitive C57BL/6N mice exposed to MNU, dietary antioxidant butylated hydroxytoluene (BHT), or both. MNU increased fetal resorptions and incidence of syndactyly, oligodactyly, polydactyly, and interdigital webbing, and decreased fetal size in both mouse strains. BHT reduced syndactyly and oligodactyly in both strains, and reduced polydactyly in C57BL/6N mice. Increased webbing in MNU and MNU+BHT groups likely represented maturational delay. Placentas from CD-1 and C57BL/6N MNU-exposed dams demonstrated decreased trophoblasts and increased necrosis of endothelium. Similar to distal limb defects, placental damage was reduced in mice receiving MNU+BHT. These results suggest that placental damage and fetal defects caused by MNU are in part ROS-mediated, and reduced distal limb defects following MNU+BHT may be related to improved placental integrity and function.

Estimated daily intakes of butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT) and tert-butyl hydroquinone (TBHQ) antioxidants in Korea.

The study was conducted to establish the estimated daily intake (EDI) of antioxidants such as butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT) and tert-butyl hydroquinone (TBHQ) in Korea. The EDIs were obtained from two sources. One of the estimations was based on the analytical determination of BHA, BHT and TBHQ in 12 food categories (ten food categories for TBHQ) and on individual dietary intake data obtained from the National Health and Nutrition Survey in 1998 (n=11 525, age > 1 year). The other EDIs of BHA, BHT and TBHQ were based on the maximum permitted levels specified in national food standards in Korea and on individual dietary intake data obtained from the National Health and Nutrition Survey in 1998 (n=11 525, age > 1 year). To establish the EDIs based on the analytical determination and on individual dietary intake data, 133 food samples in 12 food categories were selected from the foods considered to be representative sources of BHA, BHT and TBHQ in the Korean diet. Selected samples were analysed by GC with FID. BHA was not detected in any of the samples analysed. BHT and TBHQ were detected in the samples, but the levels were significantly lower than their maximum limits. The EDIs1 of BHT, and TBHQ for average consumers were 0.0156(-3), and 0.0012(-3) mg kg(-1) body weight bw day(-1) and as a proportion of the ADI were 0.0052 and 0.0002%, respectively. For 95th percentile consumers, the EDIs of BHT and TBHQ were 0.0080 and 0.0006 mg kg(-1) bw day(-1), and as a proportion of the ADI were 2.67 and 0.09%, respectively. EDIs for BHA, BHT and TBHQ based on the maximum permitted levels and on individual dietary intake data were 0.04, 0.04 and 0.04 mg kg(-1) bw day(-1), respectively. The EDIs of BHA, BHT and TBHQ for average consumers ranged from 6.00 to 14.42% of the ADI of each antioxidant. According to these results, the EDIs of BHA, BHT and TBHQ in Korea were significantly lower than ADI of these antioxidants established by the JECFA.

[Changes in composition of acid soluble proteins and DNA in chromatin of rat liver and brain bound and not bound to nuclear envelope as a function of age and under the influence of antioxidant ionol].

In two-day rat pups, the histone H1 content in the brain chromatin was higher than in the liver chromatin, as compared to histone of the nucleosome core. The H1 content in the brain chromatin decreased with the age, while in the liver chromatin it increased. At the same time, in the adult brain chromatin bound to the nuclear envelope, a high level of H1 characteristic of chromatin of the newborn rats was preserved, while in a similar chromatin of the adult liver, the H1 content increased, but still remained less than in the chromatin not bound to the nuclear envelope. In both organs, the composition and quantitation of H1 subfractions were different in chromatins bound and not bound to the nuclear envelope. The chromatin from the liver and brain bound to the nuclear envelope differed also in the composition and quantitation of minor acid soluble proteins. In the presence of the antioxidant ionol, the 5-methylcytosine content in DNA of chromatin of the rat liver bound to the nuclear envelope increased while in the chromatin not bound to the nuclear envelope, it remained unchanged. Thus the chromatins bound and not bound to the nuclear envelope differ in the composition and mount of acid soluble proteins, including histone H1, the contents of these proteins in bound and not bound chromatin are different and change with the age in different ways. The antioxidant ionol affects differently the methylation of bound and not bound chromatin.

[Activation of deoxyribonucleotide synthesis by radioprotectants and antioxidants as a key stage in formation of body resistance to DNA-damaging factors].

The responses of the systems of synthesis of deoxyribonucleotides (dNTPs), DNA, and proteins in hematopoietic organs and liver of animals to gamma-radiation, administration of radioprotectants and antioxidants as well as the dependence of these responses on the doses of radiation and drugs were studied. Radioprotectants of acute (indralin) and durable effects (indomethaphen) as well as natural (alpha2-tocopherol) and synthetic anti-oxidants (ionol or 2,6-di-tert-butyl-4-methylphenol) efficient in survival test were used. Three stages could be recognized in the standard unspecific response of the studied systems to radiation: (1) immediate increase in ribonucleotide reductase activity in the tissues within the first 30 min as a part of the integrated SOS response to DNA damage, which activates dNTP synthesis; (2) inhibition of the synthesis of dNTPs, DNA, and and (3) restoring ribonucleotide reductase activity and integral increase in the production of dNTPs, DNA, and total protein, which is essential for the development of compensatory and restorative responses of the organism. The radioprotectants significantly increased ribonucleotide reductase activity, which increased intracellular concentrations of the four dNTP types in organs during radiation exposure and three following days. Within this period, ribonucleotide reductase activity was inhibited by 40-50% in animals not treated with radioprotectants as compared to control. Balanced high pools of dNTPs in the organs of radioprotectant-treated animals provided for high-performance repair of DNA damage. The radioprotectant-induced activation of dNTP synthesis during the development of compensatory and restorative responses provides for an earlier restoration of the cellular composition and functioning of the organs. Antioxidants stimulated the synthesis of dNTPs, DNA, and proteins in animal tissues in a strict dose interval. Their effect on the studied syntheses was dose-dependent: single or multiple long-term administration of high antioxidant doses inhibited synthesis of dNTPs, DNA, and proteins. Radioprotectants and antioxidants affected the pool of blood protein Fe3+-transferrin controlling the synthesis of iron-containing ribonucleotide reductase activity in hematopoietic organs, and hence, the iron-dependent stage in DNA synthesis--dNTP synthesis. Activation of protein synthesis in organs by the studied substances increased the pools of Fe3+-transferrin and Cu2+-ceruloplasmin in the blood, which activated dNTP and DNA synthesis. Activated synthesis of dNTP, DNA, and proteins in the organs and increased pools of studied plasma proteins underlay the formation of body resistance to DNA-damaging factors.

[Anabolic effect of natural and synthetic antioxidants]. / Anabolicheskii éffekt prirodnykh i sinteticheskikh antioksidantov.

The order of responses of cell systems of organs and changes in the content of some proteins of mouse and dog blood in response to addition of natural (alpha-tocopherol) and synthetic (ionol) antioxidants was studied at the whole-body level using ERP spectroscopy, radioisotope analysis, and chemiluminescence technique. Responses were evaluated by the temporary and concentration-dependence changes in the activity of ribonucleotide reductase and the rate of protein and DNA synthesis in organs of mice, as well as by the changes in the pools of Fe3+ -transferrin and Cu2+ -ceruloplasmin in blood and the antiradical activity of blood plasma of dogs and mice. During the first 24 h of exposure to alpha-tocopherol, the activity ribonucleotide reductase in bone marrow rapidly increased, whereas the activity of this enzyme and the rate of DNA synthesis in the thymus and spleen were suppressed by 30-50% compared to the control. The changes in these parameters had a phase mode with maxima on days 2-3 and 6-8. The stimulatory effect of the antioxidant on the processes of synthesis was concentration-dependent. We found that the optimal stimulation of the synthesis of deoxyribonucleotides, DNA, and protein was achieved by single administration of alpha-tocopherol at a dose of 20 mg per dog with an average weight of 15 kg and 17 mg/kg in the case of mice. Single or repetitive administration of higher doses of alpha-tocopherol was either ineffective or even suppressed the synthesis of DNA and deoxyribonucleotides. Ionol administered at a dose of 60 mg/kg increased DNA and protein synthesis in mouse organs in 2-4 and 1.2-1.5 times, respectively, compared to the control. It was also shown that single and repetitive administration of alpha-tocopherol to dogs increased the pool of Fe3+ -transferrin and Cu2+ -ceruloplasmin in blood in 2-3 times and by 20-30%, respectively, compared to the control. It is suggested that changes in Fe3+ -transferrin pool in peripheral blood may be used for evaluation of the stimulatory effect of antioxidants on the synthesis of macromolecules in organs and for the determination of dependence of this effect on the concentration of antioxidants.

Survival and fertility of boar spermatozoa after freeze-thawing in extender supplemented with butylated hydroxytoluene.

This study evaluated the protective effect of butylated hydroxytoluene (BHT), a lipid-soluble antioxidant, against cryopreservation injuries to boar spermatozoa. In experiment 1, the lowest BHT concentrations able to reduce lipid peroxidation in boar spermatozoa were determined. Nine BHT concentrations (ranging from 0.025 to 3.2 mM) were evaluated, and the lowest (P <.05) production of malondialdehyde (MDA), as an indicator of lipid peroxidation, was obtained when BHT ranged from 0.2 to 1.6 mM. In experiment 2, sperm survivability was evaluated when BHT was added to a postthaw freezing extender by measuring the degree of sperm lipid peroxidation (using MDA production) and by measuring parameter such as motility, plasma membrane and acrosome integrity, and cell apoptosis. The ability of thawed spermatozoa to fertilize in vitro-matured oocytes and of embryos to develop to the blastocyst stage in vitro was also assessed. Pooled sperm-rich fractions collected from 3 mature Pietrain boars were frozen in 0.5-mL straws after dilution with lactose-egg yolk-glycerol-Orvus ES Paste extender supplemented with 0, 0.2, 0.4, 0.8, and 1.6 mM BHT. Postthaw sperm survival, evaluated 30 and 150 minutes after thawing, was higher in BHT-treated spermatozoa, being significant (P <.05) when the freezing extender was supplemented with 0.2, 0.4, and 0.8 mM BHT. The addition of BHT to the freezing extender resulted in a significant (P <.05) decrease in the MDA concentration in thawed spermatozoa, irrespective of the level of BHT used. BHT had no effect on oocyte cleavage rates, but the development to blastocyst was improved for embryos derived from spermatozoa frozen in extender supplemented with 0.4 mM BHT (16% vs 29% of blastocysts per total oocytes; P <.05). In conclusion, under the conditions tested in the present study, the addition of BHT to the freezing extender improved the overall efficiency of thawed boar spermatozoa.

[Effect of alpha-tocopherol and synthetic antioxidants on morpho-functional status of gonadotropic cells from the adenohypophysis of albino rats]. / Vliianie alfa-tokoferola i sinteticheskikh antioksidantov na morfofunktsional'noe sostoianie gonadotropotsitov adenogipofiza belykh krys.

Morphometric indices of gonadotropic cells obtained from adenohypophysis of white rats, both males and females, were investigated after treatment with alpha-tocopherol and synthetic antioxidants. The former stimulated the functional status of gonadotropic cells revealed in a proportional increase in both nuclear and cytoplasmic volumes. After the treatment with a synthetic antioxidant dibunolum, the volume of the cytoplasm increased in gonadotropic cells of rats of different sexes. After the treatment with a water-soluble antioxidant emoxipinum, the volume of the cytoplasm in gonadotropic cells increased only in males. The outcomes allow to consider alpha-tocopherol, in contrast to from synthetic antioxidants, as one of the modulators of the functional state of gonadotropic cells obtained from adenohypophysis.

Dietary (+)-catechin and BHT markedly increase alpha-tocopherol concentrations in rats by a tocopherol-omega-hydroxylase-independent mechanism.

The effects of dietary (+)-catechin (CAT) and BHT on plasma and tissue concentrations of alpha-tocopherol (alpha-T), gamma-tocopherol (gamma-T) and cholesterol (C) were studied in male Sprague-Dawley rats. The rats were fed the compounds during a 4-wk period at concentrations of 2 g/kg in standardized diets, low but adequate in vitamin E, with 2 g/kg cholesterol. The CAT-regimen did not affect weight gain, feed intake or organ weights. BHT did not affect feed intake but lowered the body weight and the amount of liver lipids and increased the weights of livers and lungs relative to the body weight. Rats consuming CAT had 2.5-3.5-fold increased plasma, liver and lung alpha-T concentrations, but C concentrations remained unchanged. BHT-feeding resulted in 2.4- and 1.7-fold elevation in alpha-T but approximately 50% decrease in gamma-T concentrations in blood plasma and liver, respectively. BHT also lowered total C in the liver without affecting the concentration of C in the liver lipids. To investigate whether the alpha-T-sparing action of the studied compounds was due to the inhibition of tocopherol-omega-hydroxylase, HepG2 cells were incubated with CAT or BHT in the presence of delta-tocopherol (delta-T) and the 3'- and 5'-delta-carboxychromanol metabolites in the media were analyzed by GC/MS. Neither CAT nor BHT inhibited tocopherol-omega-hydroxylase activity in hepatocyte cultures; CAT was also inactive in a rat microsomal assay. In conclusion, both dietary CAT and BHT markedly increased alpha-T concentrations in plasma and organs of Sprague-Dawley rats by a mechanism that apparently does not involve inhibition of tocopherol-omega-hydroxylase, a key enzyme in tocopherol catabolism.

Effects of dietary butylated hydroxytoluene on aflatoxin B1-relevant metabolic enzymes in turkeys.

We have shown previously that the extreme sensitivity of turkeys to aflatoxin B(1) (AFB(1)) is due to a combination of efficient AFB(1) activation by cytochrome P450s (CYPs) 1A and deficient detoxification by glutathione S-transferases (GSTs). Phenolic antioxidants such as butylated hydroxytoluene (BHT) have been shown to be chemoprotective in some animal models due, in part, to modulation of AFB(1)-relevant phase I and/or phase II activities, and we wished to determine whether BHT has a similar effect in turkeys. Ten-day-old male turkeys were maintained on diets amended with 1000 or 4000 ppm of BHT for 10 days, then sampled. Hepatic microsomal CYP 1A activity as well as conversion of AFB(1) to the putative toxic metabolite, the exo-AFB(1)-8,9-epoxide (AFBO), were significantly lower compared with control. Conversely, dietary BHT significantly increased activities of several isoforms of hepatic cytosolic GST, as well quinone oxidoreductase (QOR). Western immunoblotting confirmed that dietary BHT increased expression of homologues to rodent GST isoforms Yc1, Yc2 and Ya. There was, however, no observable BHT-related increase in GST-mediated specific conjugation with microsomally-generated AFBO. In total, our data indicates that dietary BHT modulates a variety of AFB(1)-relevant phase I and phase II enzymes, while having no measurable effect towards specific AFB(1) detoxification by GST.