Addition Of Butylated Hydroxytoluene Research Articles

THE EFFECT OF ADDITION BY BUTHYLATED HYDROXYTOLUENE (BHT) ON PHYSICAL PROPERTIES OF GEOMEMBRANE FROM RECYCLED HIGH DENSTY POLYETHYLENE (HFPE) PLASTIC WASTE

Recycled High Density Polyethylene (HDPE) has been prepared for geomembrane construction material. The aim of this research was to study the effect of Butylated Hydroxytoluene (BHT) addition as an antioxidant on the characteristics of the resulting geomembrane. Fourier Transform Infra Red (FTIR) analysis was carried out to examine functional groups of recycled HDPE and prepared geomembranes. Mechanical properties, permeability and hydrophobicity of geomembrane were observed to determined optimum BHT addition. The results of FTIR analysis on recycled HDPE showed the presence of functional groups at wavelengths 2912 and 2847 cm-1 (C-H stretching); 1474 and 1467 cm-1 (CH2 bend); 721 cm-1 (CH2 Rock). After being formulated with other materials to obtain geomembranes, it was not found new peak indicating that the presence of BHT in geomembrane only physical interaction occurs. All of geomembranes had no permeability to water and reach 100% of hydrophobicity. The highest­ tensile test value was shown by geomembrane with 0,25 b/b% of BHT which about 21.235 MPa and 16.01 MPa for before and after soil burial test for four weeks, respectively. Perhaps might be due to the interaction between BHT at low concentrations which has lower polarity and HDPE which is nonpolar has better compatibility than at other concentrations

Butylated Hydroxytoluene (BHT) Improves the Post-Thaw Semen Quality in Low-Dose Sperm Cryopreservation in Murrah Buffalo Bull

BACKGROUND: Cryopreservation is an important technique for the long-term storage of semen for artificial insemination (AI). Buffalo spermatozoa are sensitive to cryopreservation procedures because of the presence of a high amount of polyunsaturated fatty acids in the plasma membrane. OBJECTIVE: To study the effect of different concentrations of BHT on the quality of Murrah buffalo bull semen for low-dose cryopreservation. MATERIAL AND METHODS: Semen was collected from four high fertile Murrah buffalo bulls (6 ejaculates each) using an artificial vagina. A total of 24 ejaculates were collected from each bull twice a week using an artificial vagina. Every sample was split into four parts: Control without additives; and three treatments with BHT at 0.5 mM, 1 mM or 2 mM. Semen was cryopreserved at low-dose sperm cryopreservation of 20, 15, 10 and 5 million sperm per aliquot after supplementation of BHT. Semen samples were evaluated for fresh, pre-freeze and post-thaw stages. RESULTS:There was a significant increase (p<0.05) in sperm quality parameters, such as progressive motility (%), viability (%), HOST response (%), acrosome integrity (%) and post-thaw motility, with the addition of 0.5-1 mM BHT. CONCLUSION: The addition of BHT in Murrah buffalo semen improves the low dose cryopreservation quality in a dose-dependent manner.

Antioxidant, Antimicrobial and Physicochemical Properties of Beef Sausages Enriched with an Aqueous Extract of Senduduk (Melastoma malabathricum L.) Leaf

The use of natural products in sausages has become a new trend for health reasons. A natural product that could be incorporated into sausages is an extract of the senduduk (Melastoma malabathricum L.) leaf. Senduduk is an abundant shrub herb in Indonesia. This kind of plant is mostly used as a traditional medical remedy and as an ingredient in some culinary recipes. This study was carried out to investigate the effect of an aqueous extract of senduduk leaf (SLE) on the antioxidant, antimicrobial and physicochemical properties of beef sausage. Four treatments were used: ingredients consisting of beef, vegetable oil, skim milk, tapioca, salt, phosphate, ice cubic, garlic, pepper, dan nutmeg as a Control; the Control ingredients plus 0.01% of butylated hydroxytoluene (BHT); the Control ingredients plus 0.83% of SLE (SLE-1), and the Control ingredients plus 1.1% of SLE (SLE-2). All ingredients of each formula were homogenously blended and the sausage mix was cooked. The addition of BHT and SLE affected the proximate composition, with the moisture content decreasing as the duration of chilled storage increased. The addition of SLE lowered the pH and aw value and both tended to increase during chilled storage. SLE also enhanced the WHC of the sausages which increased in value during chilled storage. The addition of BHT and SLE could increase the antioxidant activity of the sausages as indicated by scavenging DPPH free radicals. SLE in sausages could inhibit microbial growth during chilled storage. It can be summarized that the addition of an aqueous extract of senduduk could improve the physicochemical, antioxidant and antimicrobial properties of beef sausages.

Accelerated Stability Test of Snakehead Fish Extract (Channa striata) and Kelulut Honey (Heterotrigona itama) Ointment Combination with BHT as an Antioxidant

The oil phase of adeps lanae was used as a basis in the ointment formulation with snakehead fish extract and kelulut honey, which influences wound healing. Adeps lanae has the potential to become rancid due to the high water content in it (25-30%). The use of oil-soluble antioxidants such as Butyl Hydroxy Toluene (BHT) can overcome the rancidity in oil. This study aimed to see how adding BHT as an antioxidant affects the ointment's stability. The ointment is prepared in three different BHT concentrations: F1 (0.0075%), F2 (0.05%), F3 (0.1%), and a control for comparison. Organoleptic tests, homogeneity, spreadability, adhesion, protection power, and acid number were used to evaluate the stability of the preparation over 28 days at 40 ± 2°C / RH 75 ± 5%. The data obtained were analyzed statistically using One Way ANOVA. The addition of BHT affected the preparation's spreadability, adhesion, and acid number but did not affect the organoleptic, homogeneity, or protective power. At F3, there is a significant difference in spreadability (5.57 ± 0.21 cm) and adhesion (89.00±2.00 seconds) based on statistical analysis. F3 is the best ointment based on the physical characteristics test results and the minimum increase in acid number (8.83 mg KOH/g).

Effects of the stability of 2-methyl furan and 2, 5 dimethyl furan on the autoignition and combustion characteristics of a gasoline-like fuel

Renewable liquid fuels have potential to greatly reduce the carbon footprint of the transportation sector while leveraging existing powertrain technologies and infrastructure. Prior studies identified a mixture of 2-methyl furan and 2, 5 dimethyl furan as one of the most promising components for formulating renewable gasoline fuel blends. Within the Co-Optima initiative, this furan mixture was used in the formulation of a custom renewable gasoline-like fuel, termed CB#2, that contains 40% of the furans in volume. Although CB#2 has demonstrated better performance than regular E10 gasoline for advanced compression ignition and boosted spark ignition engines, several concerns arose regarding stability of furans. Stability can be improved by adding antioxidant additives, but the effect of furans’ stability and the antioxidants on the ignition and combustion characteristics of gasoline fuels is unknown.In the present work, the effects of the stability of 2-methyl furan and 2, 5 dimethyl furan and the addition of butylhydroxytoluene on the autoignition and combustion characteristics of CB#2 were studied. Experimental measurements were performed using unstabilized CB#2 and CB#2 stabilized using butylhydroxytoluene (added in a concentration of 250 ppm to the furan species). From the test result, both fuel batches showed the same octane rating. Moreover, homogeneous charge compression ignition engine experiments at naturally aspirated and lean conditions showed that the two batches have the same autoignition reactivity and combustion characteristics. Therefore, fuel performance was not affected by the stability of the furans and the addition of butylhydroxytoluene at the concentration explored in this paper.

Oxidative and nitrosative stress in frozen-thawed pig spermatozoa. I: Protective effect of melatonin and butylhydroxytoluene on sperm function

The addition of antioxidants to the cryopreservation medium has been shown to exert a positive effect on the quality of frozen-thawed sperm in different species. The objective of the present study was to evaluate the effects of supplementing the freezing medium with butylhydroxytoluene (BHT) and melatonin (MEL) in frozen-thawed pig spermatozoa. With this purpose, six ejaculates coming from six separate boars were cryopreserved in traditional freezing medium (i.e. lactose/egg-yolk/glycerol; Control) supplemented with 1.0 mM BHT (BHT-1), 2.0 mM BHT (BHT-2), 0.01 μM MEL (MEL-1) and 1.0 μM MEL (MEL-2). We evaluated sperm viability, membrane lipid disorder, acrosome integrity, mitochondrial membrane potential, lipid peroxidation, oxidation of thiol groups, and levels of total reactive oxygen species (ROS), peroxynitrite and superoxide anion (·O2−). We also analysed total (TM) and progressive sperm motilities (PM), and kinetic parameters at post-thaw (T0, T30 and T60). The BHT-2 and MEL-2 groups presented higher viability and acrosome integrity, and lower levels of peroxynitrite, ·O2− and lipid peroxidation than the control (P < 0.05), whereas MEL-2 diminished the levels of total ROS (P < 0.05). TM and PM were not affected by the treatment, while, LIN and STR shows differences between experimental groups. In conclusion, the addition of BHT and MEL to cryopreservation medium diminishes oxidative and nitrosative stress markers, which has repercussions for the integrity of plasma and acrosomal membranes of frozen-thawed spermatozoa.

4-Hydroxynonenal is An Oxidative Degradation Product of Polysorbate 80.

Polysorbates (PS) are used in biopharmaceuticals to stabilize therapeutic proteins. Oxidative degradation of (poly)unsaturated fatty acids (PUFAs) contained in PS was shown to lead to α,β-unsaturated carbonyls. The n-6-PUFA linoleic acid accounts for up to 18% of all FAs contained in multi-compendial grade PS80. 4-Hydroxynonenal (HNE) is highly reactive towards nucleophilic amino acids, potentially leading to covalent protein modifications. This study tests whether HNE may be a pharmaceutically relevant PS80 peroxidation product. Since HNE was not directly detectable in the PS80 matrix by UV and MS, a new quantification method was established. After derivatization with 2,4-dinitrophenyl hydrazine (DNPH) and extraction of the formed hydrazone with a salting-out assisted liquid-liquid extraction, the HNE-DNPH adduct was analyzed by multiple reaction monitoring. Kinetic oxidation studies were conducted incubating PS80 in presence and absence of the antioxidant butylhydroxytoluene (BHT). HNE was confirmed as PS80 degradant in oxidatively stressed samples. BHT was shown to prevent its formation. HNE is a detectable PS80 degradation product raising questions about the potential impact on critical quality attributes of biopharmaceuticals formulated with PS80. Addition of BHT prevented HNE formation under oxidative stress. Consequently, BHT might be a valuable additive in PS used in biopharmaceuticals.

The investigation of the use of dried Morchella conica powder on lipid oxidation, colour, textural and sensory properties of sucuks during fermentation

The influences of dried Morchella conica powder (MCP) on quality and sensory properties of sucuks were determined. Sucuks were prepared by adding 0% (control), 1.5% MCP, 3% MCP, 0.02% butylated hydroxytoluene (BHT), and sodium nitrate/nitrite (0.01% NaNO3 /0.005% NaNO2) and lipid oxidation, colour, texture, and sensory features were analysed. Sucuks with the addition of MCP had much lower (P<0.05) peroxide values than with the addition of BHT and nitrate/ nitrite and the control samples. It was found that the L* and a* values of sucuks with the addition of 1.5% and 3.0% MCP decreased significantly (P<0.05) compared to sucuks with nitrate/nitrite and BHT addition and control samples during fermentation. Hardness and chewiness values decreased with the addition of MCP (P<0.05) during fermentation. The sucuks with MCP added at 1.5% and 3% showed the highest overall sensory acceptance. Thus, proper amount of dried MCP addition may be effective on enhancing sucuk quality properties.

Short-term storage sperm of coho salmon (Oncorhynchus kisutch) at 4 °C: Effect of sperm: Extender dilution ratios and antioxidant butyl-hydroxytoluene (BHT) on sperm function

Short-term storage of semen is a necessary key procedure in fish; it allows maximizing the use of gametes. Nevertheless, sperm quality decreases during storage has been associated with oxidative stress damage due to an increase in reactive oxygen species (ROS) during storage. This study was designed to optimize a short-term storage protocol for Coho salmon (Oncorhynchus kisutch) spermatozoa, evaluating the effect of extender dilution and the addition of butyl-hydroxytoluene (BHT) antioxidant on sperm function parameters. In the first experiment, fresh semen was diluted in Storfish®: extender dilution (1:2 and 1:3) and a control sample undiluted and stored at 4 °C for 7-days. In both experiments motility (MO), viability and integrity of plasma membrane, mitochondrial membrane potential (MMP) and superoxide anion level (O2−) were evaluated at 0, 3 and 7 days. Result shows that, 1:3 dilution maintained a higher sperm function for a longer period time. In the second experiment, spermatozoa were suspended in Storfish® (1:3) supplemented with two different concentrations of BHT (1.0 mM and 2.0 mM) and a control sample without antioxidant and stored at 4 °C for 7 days. The results demonstrated that, antioxidant-supplemented samples greater MO than control samples (P < 0.05). The viability remained >75% during storage in all groups. MMP was higher in 2.0 mM BHT compared to 1.0 mM and control (P < 0.05), in addition, this concentration reduced O2− level (P < 0.05). In conclusion, sperm: extender dilution 1:3 and adding of 2.0 mM BHT in sperm storage extender may enhance protection sperm function in Oncorhynchus kisutch against effects harmful of the oxidative stress during the in vitro storage.

Feasibility study of employing diverse antioxidants as an additive in research diesel engine running with diesel-biodiesel blends

This study examines the prospect of utilizing biodiesel from non-edible, waste seed derived from grapefruit. The prepared biodiesel was intended to replace the diesel fuel in a research diesel engine. Two antioxidants namely BHA (Butylated Hydroxyanisole) and BHT (Butylated Hydroxytoluene) are partially blended to enhance the ignition patterns of diesel/biodiesel blends. 30% volume of grape-seed biodiesel (BD) is blended with 70% volume of diesel and identified as base fuel. 10% volume of two antioxidants is equally blended to base fuel to enhance its physic-chemical properties. Upon blending the antioxidants, no change in phase was observed for base fuel. Performance results found that BHA and BHT addition lowered 1.2–1.4% of BSFC, and improved 0.7–1.1% of BTE of diesel/biodiesel blends respectively. Besides, BHA and BHT inclusion improved the physio-chemical properties of diesel/biodiesel blends and produced a prominent increase in In-Cylinder Pressure (ICP) and heat release rate (HRR). Emissions results revealed that BHA and BHT addition lowered 1.8–2.1%% of CO, 2.2–2.6% of HC of diesel/biodiesel blends. Further, a higher cetane number of BHA and BHT improved the combustion and lowered smoke (2.9–3.4%) and NO (1.9–3.7%) emission for diesel/biodiesel blends.

Stabilizing Polysorbate 20 and 80 Against Oxidative Degradation

Polysorbates are stabilizers typically required in therapeutic protein formulations. On account of their chemical structure, polysorbates are prone to degradation, which can render a pharmaceutical product instable or incompliant. The purpose of this study was to investigate if the addition of butylhydroxytoluene (BHT) and butylhydroxyanisole (BHA) protects polysorbate 20 (PS20) and polysorbate 80 (PS80) against oxidative degradation. PS20 and PS80 solutions containing BHA, BHT—or as control without an antioxidant—were stressed by exposure to air at 40°C for 7 weeks. The following assays were performed: ferrous oxidation-xylenol orange assay, liquid chromatography coupled to UV and mass spectrometry (MS), pH measurement, liquid chromatography fluorescence micelle assay, headspace–gas chromatography coupled with MS. PS20 and PS80 solutions containing an antioxidant were found to be more stable, as demonstrated by lower peroxide levels, lower free fatty acid contents, stable pH, intact polysorbate micelle structure/composition, and less volatile degradants. PS20 and PS80 solutions containing BHT or BHA are more stable against oxidative degradation compared to nonstabilized solutions. It might be beneficial to formulate bulk polysorbate with the antioxidant(s) to ensure stabilization during all process steps.

Influence of butylated hydroxytoluene addition to cryodiluents on freezability and DNA integrity of Boer and Zaraibi buck spermatozoa

Objective: To evaluate the cryoprotective effect of butylated hydroxytoluene on buck frozen semen. Methods: Semen was collected from Boer (n=6) and Zaraibi (n=6) bucks by electroejaculator for 5 weeks. Semen aliquots were diluted at 38 °C in Tris-buffer with egg yolk 15.0% (vol/vol) (Tris- egg yolk extender) or soya lecithin 2.5% (weight/vol) (Tris-soya lecithin extender) supplemented with butylated hydroxytoluene at 0.0 (as the control), 0.5, 1.0, 2.0 and 4.0 mM. Post-thawing motility (at 400x magnification), plasma (hypo-osmotic swelling test), acrosome (Trypan blue/Giemsa dual staining) membranes, DNA (comet assay), and lipid peroxidation (by malondialdehyde concentration) were assessed. Results: Spermatozoa motility was enhanced by butylated hydroxytoluene in Tris-soya lecithin extender at 0.5 mM in the two breeds, and in Tris-egg yolk extender at 1.0 mM in Boer and at 2.0 mM in Zaraibi bucks for up to 3 h post-thawing. Plasma and acrosome membranes and DNA integrity of the two breeds were maximally high with butylated hydroxytoluene at 1.0-2.0 mM in Tris-egg yolk extender and at 0.5-1.0 mM in Tris-soya lecithin extender. Lipid peroxidation was minimal with butylated hydroxytoluene at 1.0-2.0 mM in Tris-egg yolk and soya lecithin extenders in the two breeds. Butylated hydroxytoluene at 4.0 mM deteriorated spermatozoa motility, and plasma and acrosome membranes. Conclusions: The consequence of butylated hydroxytoluene on buck frozen-thawed spermatozoa varies with the levels of supplementation, buck breed, and phospholipid source in the extender. Semen parameters of Boer buck are better in their response to butylated hydroxytoluene than Zaraibi buck. Butylated hydroxytoluene at 1.0 and 2.0 mM in Tris-egg yolk extender, and at 0.5 mM in Tris-soya lecithin extender represents the best concentrations and profitably improves the semen quality of buck semen.

Direct Analysis of Synthetic Phenolic Antioxidants, and Fatty Acid Methyl Ester Stability in Biodiesel by Liquid Chromatography and High-Resolution Mass Spectrometry

Methods to identify and quantify synthetic phenolic antioxidants, 3-tert-butyl-4-hydroxyanisole (BHA), butylated hydroxytoluene (BHT), tert-butyl-hydroquinone (TBHQ) and propyl gallate (PG), in biodiesel samples by using reversed-phase liquid chromatography (LC) were developed. Using a C18 phase with LC and UV detection showed co-elution between BHT and fatty acid methyl esters (FAME) in the biodiesel sample, whereas an alkyl phenyl modified stationary phase resulted in good separation of all antioxidants from the fatty acid matrix, and allowed more accurate quantification of antioxidants in biodiesel samples. The latter column was applied for further study. Calibration curves for the four antioxidants were constructed, and the limit of detection estimated. Good calibration linearity was observed over the investigated concentration range of 10–80 ppm, with correlation coefficients (R2) ranging from 0.9986 to 0.9995 for all antioxidants. LOD values of 0.010, 0.015, 0.0125 and 0.030 ppm, and recoveries of 70 ± 2, 85 ± 2, 103 ± 2 and 92 ± 4% for PG, TBHQ, BHA and BHT at injected concentrations of 35 ppm were established, respectively. The method was applied for quantification of antioxidants in biodiesel without addition of spiked antioxidants, then for biodiesel spiked with the four antioxidants, and a commercial source of biodiesel with BHT addition. Identification of FAME in the biodiesel samples was performed by using an instrument capable of ultra-high performance LC hyphenated with an electrospray Orbitrap mass spectrometer (UHPLC–ESI-OrbitrapMS). The stability of antioxidants and FAME in different samples was then investigated. Total FAME C18 content decreased to 52 ± 4% w/w after 1 week, and 29 ± 6% w/w after 8 weeks in the test sample without antioxidants; FAME content and antioxidant composition were stable in the samples with antioxidants added, even after 8 weeks exposure to sunlight.

ADIÇÃO DE HIDROXITOLUENO BUTILADO (BHT) NO DILUIDOR ACP-106c PARA CONGELAÇÃO DE SÊMEN CANINO

Resumo: Este estudo foi realizado para determinar o efeito de hidroxitolueno butilado (BHT) sobre a qualidade do sêmen canino congelado e descongelado, utilizando o diluidor à base de água de coco em pó (ACP-106c). Para tanto, foram realizadas quinze coletas de sêmen provenientes de cinco cães. O sêmen obtido foi diluído em ACP-106c acrescido de glicerol e gema de ovo. As amostras foram então transferidas para tubos contendo diferentes concentrações de BHT (0; 0,5; 1,0 e 2,0 mM). Em seguida, as amostras foram envasadas, congeladas e armazenadas em nitrogênio líquido. O sêmen coletado foi avaliado in natura quanto aos seguintes parâmetros: coloração, volume da fração espermática, motilidade total, vigor, concentração, morfologia e funcionalidade de membrana espermática. Após uma semana, as amostras foram descongeladas e avaliadas por meio de análise computadorizada, como também foram realizadas análises da funcionalidade de membrana e da morfologia espermática. A motilidade progressiva no grupo BHT 2,0 mM foi significativamente superior (P &lt; 0,05) do que a do grupo BHT 0 mM (27,6 ± 11,7% vs. 19,0 ± 9,5%, respectivamente). Em todos os demais parâmetros avaliados, não houve diferença entre os grupos testados. Portanto, conclui-se que a adição do BHT ao diluidor ACP-106c não afetou a qualidade do sêmen canino pós-descongelação.

Effect of storage time, temperature, antioxidant and thawing on fatty acid composition of plasma, serum and red blood cells – A pilot biobank study

BackgroundIt studies on the factors that affect the stability of fatty acid profiles from human blood specimens are generally performed by evaluating the effect of a single factor on an individual fatty acid and excluding a considerable amount of data from the total fatty acid profiles. MethodsThe stability of fatty acids from plasma, serum and red blood cells (RBC) was evaluated in terms of time, temperature, antioxidant and thawing. The fatty acids were methylated and analyzed by gas chromatography. The large volume of data is evaluated simultaneously and automatically by observing an Excel-based colour scale that indicates whether the fatty acid profiles have changed significantly as a result of the storage time (0–52weeks), temperature (−20°C/−80°C), butylated hydroxytoluene (BHT) antioxidant (presence/absence) or thawing (single/multiple). ResultsFatty acids from plasma were stable at both temperatures (−20°C/−80°C) regardless of BHT. Fatty acids from serum without BHT degrades faster at −80°C than −20°C and fatty acids from RBC without BHT degrades faster at −20°C than −80°C. Addition of BHT inhibits this effect in serum and RBC. Multiple thawing of RBC without BHT demonstrated that polyunsaturated fatty acids were generally more susceptible for changes at −80°C than at −20°C while BHT prevents partially this effect. ConclusionsThis study draws attention to the importance of pre-analytical considerations when storing blood samples in biobanks and the need of careful judgments when analyzing fatty acids profiles.

An assessment of calophyllum inophyllum biodiesel fuelled diesel engine characteristics using novel antioxidant additives

In this present study, the effect of antioxidant additives with pure Calophyllum inophyllum methyl ester on the performance, combustion and emission characteristics has been investigated. New antioxidant additive namely Ethanox was added to the Calophyllum inophyllum biodiesel at concentrations of 200ppm, 500ppm and 1000ppm for oxides of nitrogen reductions and the experimental results were compared to Butylated hydroxytoluene antioxidant at same concentrations. An experimental study was done on a twin cylinder, four stroke diesel engine at a constant speed of 1500rpm with two different antioxidants, Ethanox and Butylated hydroxytoluene individually mixed with pure Calophyllum inophyllum at concentrations of 200ppm, 500ppm and 1000ppm by weight. The experimental results showed that the addition of antioxidants with Calophyllum inophyllum biodiesel produced higher brake specific fuel consumption and higher brake thermal efficiency compared to pure biodiesel. Significant reductions in oxides of nitrogen emissions were observed with Ethanox and Butylated hydroxytoluene addition with biodiesel at all concentrations compared to neat biodiesel. The reduction oxides of nitrogen emission was 12.6% for Ethanox 1000ppm and 21% for Butylated hydroxytoluene 500ppm compared to neat biodiesel. Comparable combustion characteristics were obtained by addition of Ethanox with biodiesel than Butylated hydroxytoluene antioxidant. Moreover, the addition of Ethanox and Butylated hydroxytoluene antioxidants with neat biodiesel increase the carbon monoxide, unburned hydrocarbons and smoke emission at all concentrations than biodiesel. Ethanox addition with biodiesel has shown better results compared to Butylated hydroxytoluene antioxidant.

Effect of seminal plasma and butylated hydroxytoluene (BHT) concentration on ram sperm freezability

The effect of removing seminal plasma (SP) by centrifugation before ram semen cryopreservation was assessed. Simultaneously, we studied the supplementation of different concentrations of BHT as antioxidant (0.6mM, 2mM and 5mM) on the extender in order to find an optimal concentration in enhancing a successful ram sperm cryopreservation. Therefore, fresh ejaculates from 8 rams of approximately 2 years old were collected, immediately mixed in equal quantities and split into 2 aliquots. One aliquot was diluted (1:5) in Tris-citric acid-glucose (TCG) solution and washed twice by centrifugation at 600×g for 10min, while the other aliquot was kept unwashed. Afterwards, washed and unwashed sperm were re-suspended in one of different TCG-based media with 5% glycerol and 15% powdered egg yolk (PEY) supplemented or not with BHT and/or SP according to the treatments: 1) washed sperm; 2) washed sperm+0.6mM BHT; 3) washed sperm+2mM BHT; 4) washed sperm+5mM BHT; 5) washed sperm+0.5mL of SP; 6) washed sperm+5mM BHT+0.5mL of SP; 7) unwashed sperm and 8) unwashed sperm+5mM BHT. Afterwards, sperm samples were refrigerated at 5° C for 4h before being packed in 0.25mL straws and frozen in nitrogen liquid vapors. After thawing, plasma and acrosomal membrane integrity and mitochondrial activity were determined by flow cytometry while sperm motility and morphometry by CASA (mean±SEM, n=6). Our results showed that seminal plasma removal enhanced sperm cryosurvival and the addition of BHT (5.0mM) improved most of the quality parameters of thawed washed and unwashed sperm, irrespective of the presence of SP in the extenders.

Thermo-Oxidative Stability Evaluation of Bullfrog (Rana catesbeiana Shaw) Oil.

Bullfrog oil (BO), a natural product obtained from recycling of adipose tissue from the amphibian Rana catesbeiana Shaw, has been recently evaluated as a therapeutic activity ingredient. This work aimed to evaluate the long-term and accelerated thermal oxidative stabilities of this product, which is a promising raw material for emulsion technology development. BO was extracted from amphibian adipose tissue at 70 °C with a yield of 60% ± 0.9%. Its main fatty acid compounds were oleic (30.0%) and eicosapentaenoic (17.6%) acids. Using titration techniques, BO showed peroxide, acid, iodine and saponification indices of 1.92 mEq·O2/kg, 2.95 mg·KOH/g oil, 104.2 g I2/100 g oil and 171.2 mg·KOH/g oil, respectively. In order to improve the accelerated oxidative stability of BO, synthetic antioxidants butylhydroxytoluene (BHT) and buthylhydroxyanisole (BHA) were used. The addition of BHT increased the oxidation induction time compared to the pure oil, or the oil containing BHA. From the results, the best oil-antioxidant mixture and concentration to increase the oxidative stability and allow the oil to be a stable raw material for formulation purposes was derived.

The use of butylated hydroxytoluene in cryopreservation of boar semen

The objective of the study was to determine the effect of butylated hydroxytoluene (BHT) on the quality and fertilizing capacity of frozen-thawed (FT) boar semen. Semen from five boars (36 ejaculates) was resuspended in lactose-egg yolk-glycerol extender supplemented with 0 (control), 1.0 (R1), 1.5 (R2) or 2.0 mM BHT (R3). Sperm quality was assessed based on motility (CASA; TM: total motility; PM: progressive motility), phosphatidylserine (PS) translocation across the plasma membrane (Annexin-V-FLuos Staining Kit) and DNA fragmentation (TUNEL Assay). The FT semen was also used for intrauterine artificial insemination (AI) of synchronized gilts. The fertilizing capacity of the FT semen was assessed on the basis of the gilt insemination rate and the number of morphologically normal embryos. The quality of the preimplantation embryos was determined by observing a TUNEL-positive reaction. The highest percentage of progressive motile and viable spermatozoa was noted in extender R3 (74.8 ±4.4% and 63.7 ±5.8%), as compared with the control (38.3 ±2.8% and 36.1 ±2.6%). The addition of BHT to the extender did not increase early apoptotic changes in the frozen-thawed spermatozoa with respect to the control. Irrespective of the variant of the extender, cryopreservation and thawing did not induce fragmentation in the boar spermatozoa. The highest number of morphologically normal embryos from inseminated gilts was observed in the case of semen cryopreserved in extender supplemented with 1.5 mM BHT. No significant differences were observed in DNA fragmentation in the expanded blastocysts from gilts inseminated with FT semen cryopreserved in the extenders analysed.

Effect of supplementation of butylated hydroxytoluene on post-thaw sperm viability, motility and membrane integrity of Hariana bulls.

Aim:This study was aimed to see the beneficial effect of butylated hydroxytoluene (BHT) as a semen additive of Hariana bull semen.Materials and Methods:The study was carried out in Hariana bulls. Twenty-four ejaculates from two bulls were used for this study. Each ejaculate was extended with standard glycerolated egg yolk tris extender and supplemented with BHT at two concentrations as 0.5 mM (T1) and 1.0 mM (T2). After dilution, equilibration and 24 h of cryopreservation, the samples were analyzed for progressive motility, sperm viability and membrane integrity.Results:Progressive motility, sperm viability and sperm membrane integrity were significantly (p<0.05) increased in the samples fortified with BHT as compared to the control during the process of cryopreservation and thawing. The BHT concentration of 1 mM revealed better results as compared to 0.5 mM.Conclusion:Addition of 1.0 mM BHT was found better in cryopreservation of Hariana bull semen compared to 0.5 mM BHT and control samples. The addition of BHT has improved the sperm quality by acting as an antioxidant thereby reducing the lipid peroxidation of the sperms.