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.

Application of new phenolic antioxidants for cryopreservation of sturgeon sperm.

Heterocyclic derivatives of butylated hydroxytoluene (BHT) were studied as cryoprotectants of the basic media for cryopreservation of the Russian sturgeon sperm. Rates of lipid peroxidation of sturgeon sperm before and after cryopreservation were reduced in the presence of the studied compounds, exceeding the effects of BHT and water-soluble analogue of vitamin E, trolox. The most efficient antioxidant has the effective concentration of 0.1 mM. Novel antioxidant agents as cryomedium supplements not only reduced the level of lipid peroxidation, but also enhanced the translational motility of the sperm of the Russian sturgeon after defrosting.

Cryoprotective effect of phosphorous-containing phenolic anti-oxidant for the cryopreservation of beluga sperm.

A cryoprotective effect of an addition of a new synthetic antioxidant - a representative of phosphorus-containing sterically hindered phenols is presented. The efficiency of the compound was shown to exceed the effect of lipid-soluble antioxidants butylated hydroxytoluene (BHT) and trolox in the conditions of cryopreservation of beluga sperm in the presence of the modified Stein's medium. It was shown that the level of carbonyl oxidation by-products, which can react with thiobarbituric acid (TBARS), in beluga sperm was inversely proportional to the motility time of sperm cells. The fertility of beluga sperm increased 2 times upon the addition of phosphorus-containing phenol to a modified Stein's medium. The prospects of the new antioxidant application to improve cryoresistance of beluga sperm in the conditions of cryopreservation for its efficient protection from the peroxidation processes are discussed.

[Chemical components of essential oils from Meconopsis oliverana and their antioxidant activity].

OBJECTIVE: To study the chemical components of essential oils from Meconopsis oliverana and their antioxidant activity. METHOD: The essential oil was extracted by steam distillation, and GC-MS analysis was used to identify its constituents. The OH free radical scavenging activity of the essential oils was evaluated with an enzyme mark instrument by assay of the ability of DPPH free radical scavenging. BHT was used as positive control. RESULT: Forty-seven compounds, account for 91.866% of the essential oils, were identified. The ability of scavenging OH and DPPH radicals of the essential oils is stronger than that of BHT. CONCLUSION: The main chemical constituents of the essential oils from M. oliverana are n-hexadecanoic acid (27.653%) and 6,10,14-trimethyl-2-pentadecanone (16.330%). And the essential oils showed strong antioxidant activity.

[The antiradical activity of plant extracts and healthful preventive combinations of these exrtacts with the phospholipid complex].

Using the chemiluminescence method, the effective concentration of antioxidants (AO) and its reactivity toward peroxyl radicals (ARA, the k7 constant) have been measured for 13 plant extracts. In fact all extracts demonstrated ARA higher than ionol. Larix dahurica, Hypericum perforatum, Potentilla fruticosa, Aronia melanocarpa and Rhaponticum carthamoides extracts showed the highest values of ARA. The combinations Aronia + Raponticum extracts; Larix + Hibiscus extracts; Schizandra +Aronia extracts were synergistic (the synergism effect beta of 38%, 33% and 22%). Apparently this phenomenon is the result of the synergistic interaction between compounds present in plant extracts. The Phospholipid complex--Lipoid S40, lacting any antioxidant effect alone, showed a potent synergistic effect with Aronia extract (beta3 = 60%), Silybum extract (beta3 = 41%). Clinical trials demonstrated, that combinations "Lipoid + Aronia extract", "Lipoid + Larix extract + Hibiscus extract", "Lipoid + Silybum extract", "Lipoid + Q10 + Rosa majalis extract" may be used as an additional component in the medicinal treatment, or as an individual prophylactic agent.

Hepatoprotective potential of Clitoria ternatea leaf extract against paracetamol induced damage in mice.

BACKGROUND AND AIM: Clitoria ternatea, a medicinal herb native to tropical equatorial Asia, is commonly used in folk medicine to treat various diseases. The aim of the present study is to evaluate the hepatoprotective and antioxidant activity of C. ternatea against experimentally induced liver injury. METHODS: The antioxidant property of methanolic extract (ME) of C. ternatea leaf was investigated by employing an established in vitro antioxidant assay. The hepatoprotective effect against paracetamol-induced liver toxicity in mice of ME of C. ternatea leaf was also studied. Activity was measured by monitoring the levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT) and billirubin along with histopathological analysis. RESULTS: The amount of total phenolics and flavonoids were estimated to be 358.99 ± 6.21 mg/g gallic acid equivalent and 123.75 ± 2.84 mg/g catechin equivalent, respectively. The antioxidant activity of C. ternatea leaf extract was 67.85% at a concentration of 1 mg/mL and was also concentration dependant, with an IC(50) value of 420.00 µg/mL. The results of the paracetamol-induced liver toxicity experiments showed that mice treated with the ME of C. ternatea leaf (200 mg/kg) showed a significant decrease in ALT, AST, and bilirubin levels, which were all elevated in the paracetamol group (p < 0.01). C. ternatea leaf extract therapy also protective effects against histopathological alterations. Histological studies supported the biochemical findings and a maximum improvement in the histoarchitecture was seen. CONCLUSIONS: The current study confirmed the hepatoprotective effect of C. ternatea leaf extract against the model hepatotoxicant paracetamol. The hepatoprotective action is likely related to its potent antioxidative activity.

Coenzyme Q10 production by Sphingomonas sp. ZUTE03 with novel precursors isolated from tobacco waste in a two-phase conversion system.

Coenzyme Q10 (CoQ10) is a widely used supplement in heart diseases treatment or antioxidative dietary. The microbial production of CoQ10 was enhanced by addition of solanesol and novel precursors recovered from waste tobacco. The novel precursors were separated by silica gel and identified as alpha-linolenic acid (LNA) and butylated hydroxytoluene (BHT) based on the effect on CoQ10 production and GC-MS. The effects of novel precursors on CoQ10 production by Sphingomonas sp. ZUTE03 were further evaluated in a two-phase conversion system. The precursor's combination of solanesol (70 mg/l) with BHT (30 mg/l) showed the best effect on the improvement of CoQ10 yield. A maximal CoQ10 productivity (9.5 mg l-1 h-1) was achieved after 8 h conversion, with a molar conversion rate of 92.6% and 92.4% on BHT and solanesol, respectively. The novel precursors, BHT and LNA in crude extracts from waste tobacco leaves, might become potential candidates for application in the industrial production of CoQ10 by microbes.

Antioxidant system for the preservation of vitamin A in Ultra Rice.

BACKGROUND: Ultra Rice grains are micronutrient-fortified, extruded rice grains designed to address specific nutritional deficiencies in populations where rice is a staple food. Vitamin A and some of the B vitamins, as well as iron and zinc, are target nutrients for fortification through Ultra Rice technology. Vitamin A is sensitive to degradation. Therefore, the original Ultra Rice formulations included stabilizers, some of which were not approved as food additives in all of the receiving markets. OBJECTIVE: To develop a new antioxidant system for improving vitamin A storage stability in Ultra Rice grains, while complying with international food regulations. METHODS: Ten formulations were prepared containing various combinations of hydrophilic and hydrophobic antioxidants, as well as moisture stabilizers. Accelerated vitamin A storage stability tests were conducted at 25 degrees, 35 degrees, and 45 degrees C with 70% to 100% relative humidity. RESULTS: The most stable samples contained one or more phenolic antioxidants, a water-soluble antioxidant, and stabilizing agents. The best results were obtained by using butylated hydroxyanisole (BHA) in combination with butylated hydroxytoluene (BHT) as the hydrophobic antioxidants and ascorbic acid as the hydrophilic antioxidant. Citric acid and sodium tripolyphosphate (STPP) were used to chelate metal ions and to stabilize moisture, respectively. The best formulations retained more than 85% and approximately 70% of the added vitamin A at 25 degrees and 45 degrees C, respectively, after 24 weeks storage. CONCLUSIONS: The best antioxidant system, composed of generally accepted food additives, improved vitamin A stability while reducing the price, thus greatly improving the commercial viability of Ultra Rice grains for use as a ricefortificant.

Antioxidant activity of Sanionia uncinata, a polar moss species from King George Island, Antarctica.

Antioxidant agents counter reactive oxygen species (ROS) and can be used in cosmetic and medicinal applications. The goal of this study was to evaluate the antioxidant activity of an Antarctic moss species from King George Island (Antarctica), tentatively designated as KSJ-M5. On the basis of morphological characteristics, KSJ-M5 was identified as Sanionia uncinata (Hedw.) Loeske (Amblystegiaceae). The identification was confirmed by comparing the partial sequence of the ITS (internal transcribed spacer) region with that in GenBank. The antioxidant activity of an ethanol extract of KSJ-M5 was evaluated by analyzing its reducing power, superoxide scavenging activity, ABTS [2,2'-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid)] cation scavenging activity, and DPPH (1,1-diphenyl-2-picrylhydrazyl) free-radical scavenging activity. The reducing power of 1 mg of KSJ-M5 extract was equivalent to 31.9 +/- 0.9 microg (Mean +/- SD, n = 3) of the commercial standard, BHT (butylated hydroxytoluene). IC(50) values of the KSJ-M5 extract for DPPH free-radical scavenging activity, superoxide scavenging activity, and ABTS cation scavenging activity were found as 356 +/- 26.8 microg/mL, 466.2 +/- 43.4 microg/mL, and 181.3 +/- 12.2 microg/mL, respectively. The total phenolic content in 1 mg of KSJM5 extract was equivalent to 12.7 +/- 2.7 microg of pyrocatechol. These results clearly showed that KSJ-M5 could be an important source of natural antioxidant agents for improved medicinal and cosmetic applications.

Inhibition of apoptosis facilitates necrosis induced by cisplatin in gastric cancer cells.

Although cisplatin has been shown to induce both apoptosis and necrosis in cancer cells, the potential interconnections between these modes of cell death induced by the drug remain unknown. We studied this phenomenon in gastric cancer cell lines and identified one cell line (SGC-7901) that underwent apoptosis, and another cell line (BGC-823) that primarily underwent nonapoptotic cell death, in response to cisplatin. Apoptosis in cisplatin-treated SGC-7901 cells seemed to be caspase dependent and was mediated, at least in part, by the BH3-only protein, Noxa. This was evidenced by the rapid upregulation of Noxa and inhibition of apoptosis by small interfering RNA knockdown of Noxa. Nonapoptotic cell death induced by cisplatin in BGC-823 cells was characterized by lack of DNA fragmentation, delayed externalization of phosphatidylserine, caspase independence, plasma membrane disruption, and intracellular vacuole formation, indicative of necrosis. Surprisingly, blockage of apoptosis induction by a general caspase inhibitor or by Noxa small interfering RNA in SGC-7901 failed to protect against cisplatin-induced cell death. Under such conditions, SGC-7901 cells displayed cellular features associated with necrosis. Cisplatin-induced apoptosis, thus, seems to precede necrosis when the apoptotic machinery is operative. When the apoptosis program is defective, necrotic cell death takes place as an alternative pathway leading to cell demise. Induction of different modes of cell death that are interrelated in the same cells by cisplatin has the potential to be exploited in formulating new adjuvant cancer therapies.

Monitoring of BHT-quinone and BHT-CHO in the gas of capsules of Asclepias physocarpa.

Three volatile components, namely benzoic acid ethyl ester (1), 2,6-di-tert-butyl-p-benzoquinone (BHT-quinone) (2), and 3,5-di-tert-butyl-4-hydroxybenzaldehyde (BHT-CHO) (3), were detected from the gas in the capsules of Asclepias physocarpa by means of GC/MS analysis. BHT-quinone and BHT-CHO as organic pollutants are the degradation products of the antioxidant 2,6-di-tert-butyl-4-methylphenol (BHT). Ground water, lake water and/or rain water are a source of BHT metabolites in the plant Asclepias physocarpa.

Effect of the pyridoindole antioxidant stobadine on development of experimental diabetic cataract and on lens protein oxidation in rats: comparison with vitamin E and BHT.

PURPOSE: The aim of this study was to investigate the effect of dietary supplementation with the pyridoindole antioxidant stobadine on the development of diabetic cataract in rats. The findings were compared with the effect of the natural antioxidant vitamin E and the well known phenolic synthetic antioxidant butylated hydroxytoluene. METHODS: Streptozotocin induced diabetic male Wistars rats were fed for 18 weeks a standard diet or a diet supplemented with stobadine (0.05% w/w), vitamin E (0.1% w/w), butylated hydroxytoluene (BHT, 0.4% w/w), or a mixture of stobadine (0.05% w/w) and vitamin E (0.1% w/w). The progress of cataract was monitored biweekly by ophthalmoscopic inspection. Plasma glucose and body weight were recorded regularly. At the end of the experiment, the content of free sulfhydryl and carbonyl was determined in total lens proteins and in the stobadine group plasma levels of malondialdehyde were also measured. RESULTS: Long term treatment of diabetic animals with stobadine (STB), vitamin E, or BHT led to a marked delay in the development of advanced stages of cataract. At the end of the experiment, the visual cataract score was significantly decreased in the diabetic groups treated with stobadine or BHT, while vitamin E had no significant effect. Unexpectedly, combined treatment with STB+vitamin E advanced the progression of the higher stages of cataract, though without affecting the overall visual cataract score. Neither of the antioxidants exerted an effect on the glycemic state or body weight of the animals. Biochemical analyses of eye lens proteins showed significant diminution of sulfhydryl groups and elevation of carbonyl groups in diabetic animals in comparison to healthy controls. Dietary supplementation with any of the antioxidants studied did not influence the levels of these biomarkers significantly. Nevertheless, in diabetic animals, stobadine supplementation significantly attenuated plasma levels of malondialdehyde, an index of systemic oxidative damage. CONCLUSIONS: The results are in accordance with the postulated pro-oxidant role of chronic hyperglycemia, however, the direct oxidative free radical damage of eye lens proteins does not seem to be the key mechanism effective in the development of diabetic cataract. Sugar cataractogenesis appears to be a complex process, in which multiple mechanisms may be involved, including consequences of the overt oxidative stress in diabetes (e.g., protein modifying potential of toxic aldehydes generated as byproducts of carbohydrate autoxidation and lipid peroxidation). The ability of stobadine to attenuate lipoxidation reactions in diabetes may account, at least partly, for its observed anticataract action. Mechanisms involving reduction of mitochondrial damage by stobadine are also discussed.

In vitro antioxidant properties of morphine.

Morphine is implicated in diverse functions, from development to immune modulation in the central and peripheral nervous systems. It has also been used extensively in the clinical management of pain due to its potent analgesic effect. This study was designed to evaluate the in vitro antioxidant capacity of morphine using different antioxidant tests, including total antioxidant activity, reducing power, free radical scavenging, superoxide anion radical scavenging, hydrogen peroxide scavenging and metal chelating activities. Morphine exhibited strong total antioxidant activity. The concentrations of 25, 50 and 75 microgml(-1) of morphine showed 79.1, 84.3 and 92.3% inhibition on peroxidation of linoleic acid emulsion, respectively. On the other hand, at 75 microgml(-1) concentration of standard antioxidant, such as butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT) and alpha-tocopherol, exhibited 88.7, 94.5 and 70.4% inhibition on peroxidation of linoleic acid emulsion, respectively. In addition, morphine had effective reducing power, free radical scavenging, superoxide anion radical scavenging, hydrogen peroxide scavenging and metal chelating activities at the same concentrations (25, 50 and 75 microgml(-1)). These various antioxidant activities were compared to standard antioxidants such as BHA, BHT and alpha-tocopherol.

Some natural and synthetic antioxidants as stabilizers of beta-carotene conversion into vitamin A.

The effect of antioxidants (vitamins C and E, quercetin, probucol, butylated hydroxytoluene) on the oxidation of beta-carotene and its conversion into retinal under the influence of beta-carotene 15,15'- dioxygenase (CDO) from rat intestinal mucosa was studied. The activity of CDO decreased in the presence of oxidants. Antioxidants protected both the substrate and the enzyme. The extent of the protection depended on the antioxidant type. The combined injection of antioxidants and beta-carotene to animals completely or partially prevented the inhibition of the intestinal CDO which was caused by products of non-enzymatic oxidation of beta-carotene. Vitamins C and E, which protected the enzyme--substrate complex in vivo and in vitro, were found to be the most efficient protectors of beta-carotene conversion into retinal.

The application of antioxidants in investigations and optimization of photochemotherapy.

Psoralens (furocoumarins) are photosensitizers of plant origin. They are used in combination with near-ultraviolet (320-400 nm) light for the treatment of vitiligo, psoriasis, cutaneous T-cell lymphoma (CTCL), alopecia areata, eczema, and other skin diseases. Photobiological effects of psoralens in humans are numerous. They photosensitize erythema, hyperpigmentation, and skin aging, affect immune system, etc. Molecular mechanisms of photochemical reactions of psoralens with substrates are also numerous. The molecular basis and the relationships between different photobiological effects of psoralens remain the subject of discussion. The use of antioxidants is an essential instrument for clearing up these problems as well as for the progress of photochemotherapy. It was found that antioxidants (alpha-tocopherol, butylated hydroxytoluene, etc.) selectively inhibited the photochemical stage of erythema and hyperpigmentation but had no impact on the post-irradiation stages of these processes. Evidently, the basis of these processes is the reaction of psoralen-photosensitized oxidation of unsaturated lipids and the impairment of barrier functions of biomembranes, since the photochemical stage of these reactions is inhibited by the antioxidant. At the same time, antioxidants did not inhibit the therapeutic effect in the cases of psoriasis and CTCL. Thus, antioxidants can be used as a tool for improvement of psoralen photochemotherapy.

Free-radical and cyclooxygenase-catalyzed lipid peroxidation in membranes of blood cells under UV irradiation.

The data on the role of lipid peroxidation in the effects of UV irradiation of blood are reviewed. Lipid photoperoxidation in blood cells is the result of photochemical transformation of lipid hydroperoxides, both existing and newly formed ones, into free radicals and direct photolysis of photooxidants. Dark lipid autoperoxidation is also induced by UV radiation. Both peroxidation and photooxidation of lipids are inhibited by low concentrations of antiradical antioxidants. The cyclooxygenase-catalyzed peroxidation of arachidonic acid in blood cells is stimulated by UV radiation. This process is suppressed by acetylsalicylic acid and indomethacin. The therapeutic activity of the blood that was UV-irradiated and then infused into rats with peritonitis was due to the cyclooxygenase activation.

Formation of oxysterols during oxidation of low density lipoprotein by peroxynitrite, myoglobin, and copper.

Oxidation of low density lipoprotein (LDL) in the artery wall leads to the formation of cholesterol oxidation products that may result in cytotoxicity. Different mechanisms could contribute to LDL oxidation in vivo resulting in characteristic and specific modification of the cholesterol molecule. Alternatively, attack on cholesterol by chain propagating peroxyl radicals could result in the same distribution of oxidation products irrespective of the initial pro-oxidant mechanism. To distinguish between these possibilities we have monitored the formation of nine oxysterols during LDL oxidation, promoted by copper, myoglobin, peroxynitrite, or azo bis amidino propane. Regardless of the oxidant used, the pattern of oxysterol formation was essentially the same. The yields of products identified decreased in the order 7-oxocholesterol > 7 beta-hydroxycholesterol > 7 alpha-hydroxycholesterol > 5,6 beta-epoxycholesterol > 5,6 alpha-epoxycholesterol except in the case of peroxynitrite in which case a higher yield of 5, 6 beta-epoxycholesterol relative to 7-oxocholesterol was found. No formation of cholestane 3 beta, 5 alpha, 6 beta-triol, or the 24-,25-,27-hydroxycholesterols was seen. Concentration of 7-oxocholesterol levels in LDL was positively correlated with the degree of protein modification. Endogenous alpha-tocopherol in LDL or supplementation with butylated hydroxytoluene prevented oxysterol formation. Taken together these data indicate that the oxidation of cholesterol and protein in LDL occur as secondary oxidation events consequent on the attack of fatty acid peroxyl/alkoxyl radicals on the 7-position of cholesterol, and with amino acids on apoB. Furthermore, oxidant processes with atherogenic potential, such as peroxynitrite, copper, and myoglobin are capable of producing oxidized LDL containing cytotoxic mediators.