HPLC determination of chlorate metabolism in bovine ruminal fluid

Salmonella and Escherichia coli are two bacteria that are important causes of human and animal disease worldwide. Chlorate is converted in the cell to chlorite, which is lethal to these bacteria. An HPLC procedure was developed for the rapid analysis of chlorate (ClO₃ ^?), nitrate (NO₃ ^?), and nitrite (NO₂ ^?) ions in bovine ruminal fluid samples. Standard curves for chlorite, nitrite, nitrate, and chlorate were well defined linear curves with R² values of 0.99846, 0.99106, 0.99854, and 0.99138, respectively. Concentrations of chlorite could not be accurately determined in bovine ruminal fluid because chlorite reacts with or binds a component(s) or is reduced to chloride in bovine ruminal fluid resulting in low chlorite measurements. A standard curve ranging from 25 to 150 ppm ClO₃ ^? ion was used to measure chlorate fortified into ruminal fluid. The concentration of chlorate was more rapidly lowered (P < 0.01) under anaerobic compared to aerobic incubation conditions. Chlorate alone or chlorate supplemented with the reductants sodium lactate or glycerol were bactericidal in anaerobic incubations. In anaerobic culture, the addition of sodium formate to chlorate-fortified ruminal fluid appeared to decrease chlorate concentrations; however, formate also appeared to moderate the bactericidal effect of chlorate against E. coli. Addition of the reductants, glycerol or lactate, to chlorate-fortified ruminal fluid did not increase the killing of E. coli at 24 h, but may be useful when the reducing equivalents are limiting as in waste holding reservoirs or composting systems required for intense animal production.     

A simple HPLC method for determination of permethrin residues in wine

An isocratic High Performance Liquid Chromatographic (HPLC) method was optimized for 3-phenoxybenzyl (1RS)-cis-trans-3-(2,2-dichlorovinyl)-2,2-dimethyl-cyclopropanecarboxylate (permethrin) residues identification and quantification in wine matrix. Analytical reverse phase (RP) C-18 column was used (25 cm × 4 mm i.d., 5 μ m) with mobile phase consisting of acetonitrile and water in ratio 70 %/30 % (v v^?1), flow-rate 2.0 mL min^?1, UV-detection at 215 nm and controlled oven temperature at 25°C. The peaks of isomers were identified with the retention times as compared to standard cis-/trans- mixture and confirmed with characteristic spectra using photodiode array detector. Under these conditions, permethrin isomers were well separated with resolution 2.8 and no interference with the naturally present wine compounds was observed. The method was validated for linearity, precision, accuracy, limit of detection (LOD) and limit of quantification (LOQ). Linear regression analysis data proved a good linear relationship (correlation coefficients, r², for cis- and trans-isomer are: 0.9995 and 0.9997, respectively) between response of the detector and concentration of permethrin isomers over a wide concentration range for both isomers (0.55 mg L^?1 ?4.40 mg L^?1). Experimental data showed mean recoveries between 93.95% and 96.58% with RSD values in range: 0.89% ?3.69%. The effect of ethanol content in the solvent on permethrin isomers peak areas was also studied and 60% v v^?1 ethanol was found to be optimal for sample preparation. The method was successfully tested on 20 commercial wine samples from the market in which no permethrin was detected. Thus, it was proved that it is suitable for routine permethrin residues analysis. The proposed method is suitable for routine analysis because of the simple sample preparation, acceptable run-time, low cost and its applicability with conventional instruments.     

Effects of pulp mill chlorate on Baltic Sea algae

The long-term effects of pulp mill chlorate on different algal species of the Baltic Sea were studied in land-based model ecosystems simulating the littoral zone. Brown algae (Phaeophyta) exhibited an extraordinarily high sensitivity to chlorate and pulp mill effluents containing chlorate. All brown algal species ceased growth or showed major signs of toxicity at all concentrations tested, down to microgram per litre levels. EC50 levels for growth of Fucus vesiculosus were about 80-100 microg ClO3- litre(-1). Blue-green algae (Cyanophyta) were not deleteriously affected nor were green algae (Chlorophyta). The perennial and annual species of red algae (Rhodophyta) were also unaffected by the effluents. Diatoms did not show any sensitivity and phytoplankton (fresh- and brackish water) were particularly insensitive. A phanerogam, Zostera marina was also unaffected by the treatments.     

Comparison of supercritical fluid extraction and Soxhlet extraction for the determination of PCBs in seaweed samples

The efficiency of supercritical fluid extraction for the determination of 12 polychlorinated biphenyls from algae samples is compared to Soxhlet extraction. Analytical detection limits for the individual congeners ranged from 0.62 microgl(-1) to 19 microgl(-1). Recovery was tested for both methods using standard addition procedure. At maximum spike level of concentration, the mean recoveries were not significantly different (P>0.05) of all PCBs studied, with the exception of PCBs 28, 52, 77 and 169. Method precision for Soxhlet extraction (< or =3.9%) was slightly better than for SFE (< or =9.2%). Although both methods yield comparable results, SFE offers the advantage of detecting all PCBs studied at lower concentrations, reducing extraction time, and reducing the amount of solvents needed. The optimized methods were applied to the analysis of three real seaweed samples, except for PCB101 the concentrations of all PCBs were low or below the detection limits. The levels of PCB101 found in sample 1 were 6.6+/-0.54 ng g(-1) d.w., in sample 2 the levels were 8.2+/-0.86 ng g(-1) d.w. and in sample 3 they were 7.7+/-0.08 ng g(-1) d.w.     

Dietary fiber content influences soluble carbohydrate levels in ruminal fluids

The soluble carbohydrate concentration of ruminal fluid, as affected by dietary forage content (DFC) and/or ruminally undegradable intake protein content (UIPC), was determined. Four ruminally cannulated steers, in a 4 × 4 Latin square design, were offered diets containing high (75 % of DM) or low (25 % of DM) DFC and high (6 % of DM) or low (5 % of DM) UIPC, in a 2 × 2 factorial arrangement. Zinc-treated SBM was the primary UIP source. Soluble hexose concentration (145.1 μM) in ruminal fluid (RF) of steers fed low DFC diets exhibited a higher trend (P = 0.08) than that (124.5 μM) of steers fed high DFC diets. UIPC did not modulate (P = 0.54) ruminal soluble hexose concentrations. Regardless of diet, soluble hexose concentration declined immediately after feeding and did not rise until 3 h after feeding (P < 0.0001). Cellobiose (≈90 %) and glucose (≈10 %) were the major soluble hexoses present in RF. Maltose was not detected. Soluble glucose concentration (13.0 μM) was not modified by either UIPC (P = 0.40) nor DFC (P = 0.61). However, a DFC by post-prandial time interaction was detected (P = 0.02). Pentose concentrations were greater (P = 0.02) in RF of steers fed high DFC (100.2 μM) than steers fed low DFC (177.0 μM). UIPC did not influence (P = 0.35) soluble pentose concentration. The identity of soluble pentoses in ruminal fluid could not be determined. However, unsubstituted xylose and arabinose were excluded. These data indicate that: (i) soluble carbohydrate concentrations remain in ruminal fluid during digestion and fermentation; (ii) slight diurnal changes began after feeding; (iii) DFC influences the soluble carbohydrate concentration in RF; and (iv) UIPC of these diets does not affect the soluble carbohydrate concentration of RF.     

Procedures of trophic chain samples preparation for determination of triazines by HPLC and metals by ICP-AES methods

The aim of this research was monitoring the distribution of atrazine and simazine as well as metals Pb, Cd, Zn, Al, Co, Ni, and V along with trophic chains: soil-vegetables and soil, carrot or grass and meat. Different techniques of herbicides extraction by means of many solvents were examined. Triazines were analysed by means of HPLC, metals by means of ICP-AES. Detection limits: LOD=0.2 microg ml(-1), determination limits: LOQ=0.73 microg ml(-1) for atrazine and LOD=0.3 microg ml(-1), LOQ=1.12 microg ml(-1) for simazine were obtained. The content (microg g(-1)) of simazine in soil was in range: 3.45-8.60, in vegetable roots: 6.62-38.15, in vegetable leaves: 2.45-31.71, in rabbit fat: 0.13-49.90. The content (microg g(-1)) of atrazine in soils was in range: 11.9-13.03, in vegetable roots: 13.61-92.90. In analysed material the particular metals after microwave or dry digestion were determined in range (microg g(-1)): Pb: 6.48-43.18; Cd: 0.11-0.57; Zn: 8.79-51.90; Al: 10.22-24.48; Co: 0.18-3.89; Ni: 0.37-6.36; V: 0.29-1.48.     

Quantitative determination of 1 -hydroxypyrene in bovine urine samples using high-performance liquid chromatography with fluorescence and mass spectrometric detection

An analytical method was developed to determine quantitatively 1-hydroxypyrene (OHP) in bovine urine samples. The procedure includes an enzymatic hydrolysis to cleave the conjugated metabolite, an enrichment step using solid phase extraction with a non-polar rinse step and elution with dichloromethane. A final clean-up on silicagel was performed before high-performance liquid chromatography (HPLC) analysis and fluorescence detection. Alternatively, HPLC and electrospray ionization in the negative ion mode applying selective ion monitoring acquisition revealed to be a highly sensitive detection method allowing the quantitation of low pg of OHP in the urine samples. The method was successfully applied to the determination of OHP in bovine urine samples from animals living in urban and rural areas. Urine concentrations of OHP were significantly higher (median 8.6 microg l(-1)) of bovines living close to a highway.     

Ethoxyresorufin-O-deethylase (EROD) activity and bile metabolites as contamination indicators in Baltic Sea perch: determination by HPLC

Exposure of Baltic Sea perch (Perca fluviatilis) to organic contaminants was investigated by means of liver enzyme activity, measured as ethoxyresorufin-O-deethylase (EROD) activity, and PAH metabolites excreted to bile. Female perch used in the study were caught in coastal waters of the western Gulf of Finland. Reaction conditions were optimised for determination of EROD activity in liver. A high-performance liquid chromatography (HPLC) method with fluorescence detection was developed to separate the interfering fluorescent signal from the signal due to EROD activity. EROD activity in perch varied within a concentration range of 0.30-14 pmol min(-1)mg(-1) protein. Recent PAH exposure was indicated in enhanced levels of the bile metabolite 1-hydroxypyrene, which varied between 213 and 1149 microg kg(-1). No correlation was indicated between hepatic EROD activity and concentration of 1-hydroxypyrene.     

Interaction of tebuconazole with bovine serum albumin: determination of the binding mechanism and binding site by spectroscopic methods

Abstract

This study investigates the interaction between tebuconazole and bovine serum albumin (BSA) in a physiological buffer (pH = 7.4) using the fluorescence quenching method to obtain the apparent binding constants (K) and number of binding sites (n) in the interaction between tebuconazole and BSA. The results revealed that tebuconazole can quench the intrinsic fluorescence of BSA through a static quenching procedure. It also shows that the thermodynamic parameters of enthalpy change (ΔH) and entropy change (ΔS) are negative, indicating that the interaction of tebuconazole with BSA is mainly driven by van der Waals forces and hydrogen bonds. The process of binding was a spontaneous process in which Gibbs free energy change was negative. The distance of r between the donor (BSA) and acceptor (tebuconazole) was calculated to be 0.68?nm based on Forster’s non-radiative energy transfer theory. Analysis of synchronous fluorescence, three-dimensional fluorescence and circular dichroism (CD) spectra demonstrates that tebuconazole can induce conformational changes of BSA.     

Multicriteria optimisation of a simultaneous supercritical fluid extraction and clean-up procedure for the determination of persistent organohalogenated pollutants in aquaculture samples

A useful tool based on a single-step extraction and clean-up procedure for the determination of 15 organohalogenated pollutants (including brominated flame retardants) in aquaculture samples, using aluminium oxide basic and acidic silica gel in the supercritical extraction cell followed by gas chromatography with electron capture detection or mass spectrometry has been developed. This effective clean-up step ensures a minimum of chromatographic difficulties related to complex matrix components such as aquaculture feed. The extraction procedure has been screened by a fractional factorial design for the preliminary statistically significant parameters. The factors selected were extraction temperature, pressure, static extraction time, dynamic extraction time and carbon dioxide flow rate. The Doehlert design, followed by a multicriteria decision-making strategy, was then performed in order to determine the optimum conditions for the two most significant factors: pressure (165 bar) and dynamic extraction time (27 min). Under optimal conditions, the procedure developed with GC-MS/MS provides an excellent linearity, detection (0.01-0.2 ng g(-1)) and quantification limits (0.05-0.8 ng g(-1)) for most of the analytes investigated. The feasibility of the proposed supercritical fluid extraction method was validated by analysing two reference materials and fish feed and shellfish samples with satisfactory results.     

Evaluation and validation of two different chromatographic methods (HPLC and LC-MS/MS) for the determination and confirmation of ochratoxin A in pig tissues

This study was undertaken to compare two different analytical methods for the determination and confirmation of ochratoxin A (OTA) in blood serum, kidney and liver of pigs. Sample clean-up was based on liquid-liquid phase extraction. The detection of OTA was accomplished with high-performance liquid chromatography (HPLC) combined either with fluorescence detection (FL) or electro spray ionization (ESI+) tandem mass spectrometry (MS–MS). Comparative method evaluation was based on the investigation of 90 samples of blood serum, kidney and liver per animal originating from different regions of Serbia. The analytical results are discussed in view of the respective method validation data and the corresponding experimental protocols. In general, analytical data obtained with (LC–MS–MS) liquid chromatography electro spray tandem mass spectro metry detection offered comparable good results at the sub-ppb concentration level. The results indicate that the liquid chromatography electro spray tandem mass spectrometric (LC-MS/MS) method was more specific and sensitive for the analysis and confirmation of ochratoxin A in pig tissues then high pressure liquid chromatography (HPLC) method after methylation of OTA.     

西兰花中吡蚜酮残留量的高效液相色谱检测与条件优化研究

采用正交试验,优化建立了西兰花中吡蚜酮残留量的高效液相色谱分析方法.西兰花经乙腈和二氯甲烷提取,柱层析净化后,最佳检测条件为柱温20℃、流动相为乙腈-水(体积比为85:15)、进样量10μL、流速1 mL/min.该方法对毗蚜酮的最小检出量为1.0×10-10g,最低检出质量浓度为0.005 mg/kg,西兰花平均添加回收率为87.29%～93.60%,符合农药残留检测要求.     

Ion chromatographic determination of anions in rainwater

High acidity rainwater has recently become an issue in environmental pollution because of its great influence on ecological systems with special reference to the human environment. To solve this problem, the chemical behaviors of various pollutants in the air, particularly in the wet atmosphere must be studied. Ion chromatography was used for rapid analysis of trace amounts of anions.As a result, it was found that a combination of 2mM Na₂CO₃ and 5mM NaOH as an eluent was suitable for the separation of various anions. The reproducibility of determination of anions in standard solutions was found to be satisfactory with a coefficient of variation (CV) of 2–4% for F^?, Br^?, NO^?₃, SO^2?₃, SO^2?₄, and PO^3?₄, but not for NO^?₂. The determination of NO^?₂ was accomplished by preparing an eluent having almost the same composition and cencentration as that of the standard and sample solutions. The precision of the method with rainwater samples was less than 10%, and was considered to be usable for data analysis.In addition, a considerable number of ions in rainwater samples could be determined by using 1–3 ml of the sample without pretreatment. It was found that ion chromatography was one of the best means to obtain information on the chemical behaviors of trace amounts of anions in rainwater.     

水中苦味酸的测定

采用GC-ECD法,通过测定水中苦昧酸氯化生成氯化苦的量,来确定苦昧酸的浓度。利用GC-MS确定了氯化苦的出峰保留时间,研究了反应时间、次氯酸钠加入量及其他硝基芳烃化合物对苦昧酸测定的影响,进一步优化了水中苦昧酸测定的条件。应用GC-ECD法测定了太湖水中的苦味酸,较好地满足了水质检测的要求。     

Determination and characterization of chloroguaiacol conjugates in fish bile by HPLC

In this study a small-scale technique for direct analysis of metabolic conjugates of 4,5,6-trichloro- (CG-3) and tetrachloroguaiacols (CG-4) in fish bile by RP-HPLC is presented. Only one metabolite, glucuronic acid conjugate, was demonstrated in two Lake Baikal fishes (Leuciscus leuciscus baikalensis and Cottus kessleri) exposed to CG-3 or CG-4 at 6 degrees C for 1-2 days. In Leuciscus the ratio between free CG-4 in the ambient water and conjugated CG-4 in the bile averaged 264,000. Intraperitoneal injections of tetrachloroveratrol (CV-4) into Thymallus arcticus baikalensis did not reveal any traces of free CV-4 or CG-4 glucuronide in the bile in 2 days at 5 degrees C. We therefore suggest that CV-4 was not demethylated and subsequently conjugated with glucuronic acid in the liver of this fish species.     

高效液相色谱法测定南昌市环境空气PM10中16种多环芳烃

建立了以二极管阵列检测器和荧光检测器串联的高效液相色谱分析方法,在标样未完全分离的情况下,采用双激发波长有效地改善了色谱分离条件.在设定的色谱条件下,各种多环芳烃(PAHs)的检出限为0.11～39.83μg/L,平均回收率为76.7%～98.3%,相对标准偏差为3.6%～12.6%.在南昌市布设4个环境空气采样点,测定PM10中PAHs含量.结果表明,八一广场、南昌市区二中老校区和罗家集区苯并(a)芘日均质量浓度最大值均超过<环境空气质量标准>(GB 3095-1996)的限值,PAHs污染状况较严重.     

The metabolism of 2,3,7,8-tetrachlorodibenzofuran in the rat

Single doses of 2,3,7,8-tetrachlorodibenzofuran (2378-TCDF) were administered orally to bile duct cannulated rats and bile collected for up to 48 hours. The bile was analysed for the parent compound and metabolites. In organic extracts, which had been methylated and purified by TLC, 13 chlorinated metabolites were detected that were not present in the bile of untreated animals. Four of these, tetrachloromethoxy-, trichloromethoxy-and two trichlorodimethoxy-dibenzofuran isomers were present in major amounts. The remaining compounds, found only in minor or trace amounts very likely originate from other chlorinated dibenzofurans present as impurities in 2378-TCDF. No parent compound was detected in these samples.     

Effects of gradually increasing drying temperatures on energy aspects,fatty acids,chemical composition,and in vitro ruminal fermentation of acorn

Environmental Science and Pollution Research - Acorns are commonly used to meet energy, protein, and mineral needs of livestock in various parts of the world. However, since acorns have quite a...     

Rat metabolism of polychlorinated dibenzo-p-dioxins

In rats, dibenzo-p-dioxin, 1-chlorodibenzo-p-dioxin, 2-chlorodibenzo-p-dioxin, 2,3-dichlorodibenzo-p-dioxin, 2,7-dichlorodibenzo-p-dioxin, 1,2,4-trichlorodibenzo-p-dioxin and 1,2,3,4-tetrachlorodibenzo-p-dioxin are metabolized to mono- and dihydroxy derivatives, whilst in case of dibenzo-p-dioxin and both the two monochloro isomers, also sulphur containing metabolites are excreted. Primary hydroxylation exclusively takes place at the 2-, 3-, 7- or 8-position in the molecule. In none of the experiments metabolites resulting from fission of the C-O bonds (ortho, ortho'-dihydroxychlorodiphenyl ethers, chlorocatechols) or hydroxylated derivatives thereof, were detected. No metabolites were found from octachlorodibenzo-p-dioxin.     

Method development for determination of fluroxypyr in soil

Four methods were developed for the analysis of fluroxypyr in soil samples from oil palm plantations. The first method involved the extraction of the herbicide with 0.05 M NaOH in methanol followed by purification using acid base partition. The concentrated material was subjected to derivatization and then cleaning process using a florisil column and finally analyzed by gas chromatography (GC) equipped with electron capture detector (ECD). By this method, the recovery of fluroxypyr from the spiked soil ranged from 70 to 104% with the minimum detection limit at 5 microg/kg. The second method involved solid liquid extraction of fluroxypyr using a horizontal shaker followed by quantification using high performance liquid chromatography (HPLC) equipped with UV detector. The recovery of fluroxypyr using this method, ranged from 80 to 120% when the soil was spiked with fluroxypyr at 0.1-0.2 microg/g soil. In the third method, the recovery of fluroxypyr was determined by solid liquid extraction using an ultrasonic bath. The recovery of fluroxypyr at spiking levels of 4-50 microg/L ranged from 88 to 98% with relative standard deviations of 3.0-5.8% with a minimum detection limit of 4 microg/kg. In the fourth method, fluroxypyr was extracted using the solid liquid extraction method followed by the cleaning up step with OASIS HLB (polyvinyl dibenzene). The recovery of fluroxypyr was between 91 and 95% with relative standard deviations of 4.2-6.2%, respectively. The limit of detection in method 4 was further improved to 1 pg/kg. When the weight of soil used was increased 4 fold, the recovery of fluroxypyr at spiking level of 1-50 microg/kg ranged from 82-107% with relative standard deviations of 0.5-4.7%.