Analysis Of Methanol Research Articles

In-syringe homogeneous liquid-phase microextraction followed by filtration-based phase separation for on-site extraction of chloroanilines from water samples.

This study introduces a new in-syringe homogeneous liquid-phase microextraction method for the rapid on-site extraction of chloroanilines from water samples. Extraction was performed using a plastic syringe, eliminating the use of any electrical power source. Di-(2-ethylhexyl) phosphoric acid (DEHPA) served as the extractant. The process initially involved dissolving DEHPA in an alkaline solution to obtain a homogeneous solution. Subsequently, the sodium salt of DEHPA was precipitated by salting-out, and the resulting heterogeneous mixture was filtered using a syringe filter. The precipitate containing the analytes was then dissolved in methanol for analysis by high-performance liquid chromatography. Under optimal conditions, extraction recovery for chloroanilines ranged from 26% to 71%. Method linearity was evaluated within a concentration range of 1.0-100µg/L, resulting in coefficients of determination exceeding 0.9987 for all analytes. Method detection limits ranged from 0.28 to 0.41µg/L. Intra and inter-day precision values were below 9.5% and 10.8%, respectively. The developed method was applied to determine chloroanilines in real waters, yielding acceptable recoveries ranging from 80% to 109% for spiked tap, rain, and stream waters. Additionally, the method was successfully employed for on-site extraction of target contaminants, demonstrating no statistically significant differences compared to laboratory results.

QUALITATIVE ESTIMATION OF PHYTOCHEMICALS IN DIFFERENT OCIMUM SPECIES OF CHHATTISGARH

Medicinal plants and herbs have been proved to be of great importance to the health of theindividuals and communities. The potential for developing antibacterial from higher plants appearsrewarding as it will result to the development of a phytomedicine to act against microbes. Phytochemicalscreening of Tulsi plant (Ocimum sp.) was studied in the present investigation. Phytochemical analysis ofmethanol and aqueous extracts of O. tenuiflorum, O. gratissumand O. sanctum demonstrated the presenceof phytoconstituents like Tannins, Alkaloids, Saponins, Glycosides, Terpenoids, Flavonoids, Phenols,Carbohydrates, Steroids, and Anthraquinones. Presence of such phyto constituents will increase interestwith the mankind to develop anti-microbial drugs using different species of Oscimum for varied microbes.

Standardization of extract mixture of Chaenomeles sinensis and Phyllostachys bambusoides for anti-obesity by HPLC-UV.

JM-101 is a developed functional food formula using water extract of Chaenomeles sinensis and Phyllostachys bambusoides for anti-obesity. Standardization and quality control of herb mixture is more difficult than those of single herb. Additionally, the estimation of mixing ratio is an essential requirement for standardization. This study aimed to develop an efficient analytical method for the standardization of JM-101 based on C. sinensis and P. bambusoides. Protocatechuic acid and p-coumaric acid were selected as marker compounds of JM-101. A mixture of the two medicinal materials (1:1 w/w) was extracted by water and then liquid-liquid extracted (LLE) by ethyl acetate. The supernatant was evaporated to dryness and dissolved in methanol for analysis. The extraction time, material-to-water ratio and ethyl acetate-to-water ratio were optimized by multi-response optimization based on response surface methodology (RSM). The established methods were validated in terms of linearity, precision, accuracy, repeatability, stability and recovery. The novel method based on LLE and RSM provides a sensitive, accurate analysis and excellent extraction efficiency of marker compounds in JM-101, without interruption of other compounds in JM-101. In conclusion, the developed simultaneous analytical method contributes to the standardization of two materials (C. sinensis and P. bambusoides) and JM-101.

Matrix solid-phase dispersion combined to liquid chromatography–tandem mass spectrometry for the determination of paraben preservatives in mollusks

A method for the extraction and determination of seven parabens, esters of 4-hydroxybenzoic acid, widely used as preservatives in personal care products, pharmaceuticals, etc., and two chlorinated derivatives (mono- and di-chloro methyl paraben) from mollusk samples was developed by combining matrix solid-phase dispersion (MSPD) and liquid chromatography–tandem mass spectrometry. MSPD parameters, such as solvent, solid support and clean-up sorbent, were optimized. Besides, since blank problems were observed for some parabens, these were investigated and blanks were tackled by precleaning all sorbents prior to use. Under final conditions, 0.5g of freeze-dried mollusk were dispersed with 1.2g of silica and packed into a cartridge containing 3g of C18, as on-line clean-up sorbent. This cartridge was eluted with 10mL of acetonitrile, evaporated and reconstituted in methanol for analysis. In the validation stage, successful linearity (R2>0.999), recoveries (between 71 and 117% for most analytes), precision (RSD lower than 21%) and limits of detection and quantification (LOD and LOQ, lower than 0.4 and 1.4ngg−1 dry weight respectively) levels were achieved. Finally, the new methodology was applied to mussel, clam and cockle samples. Methyl paraben was above the LOQ in five of the six samples (not found in one clam sample) at concentrations up to 7ngg−1 dry weight. Ethyl paraben was found above the LOQ in mussel and cockle samples at a concentration level around 0.3ngg−1. n-Propyl paraben was only above the LOQ in one mussel sample.

Identification of methylated avenanthramides in human plasma

Avenanthramides (AVs) are phytochemicals unique to oats consisting of an anthranilic acid and hydroxycinnamic acid linked by a pseudo‐peptide bond. We have demonstrated AV bioavailability to be low in hamsters and humans, suggesting they may be metabolized by phase II enzymes to facilitate their rapid excretion. Thus, we sought to identify the presence of phase II metabolites of 6 dominant AV isomers (A, B, C, O, P, and Q). AV‐A, ‐B, and ‐C differ in the moiety at position 3 (A: ‐H; B: ‐CH3O; and C: ‐OH) of the cinnamic acid ring, and AV‐O, ‐P, and ‐Q correspond to A, B, and C in terms of position 3 but differ in the length of the pseudo‐peptide linkage. We conducted a placebo‐controlled, crossover study in 10 healthy adults (60.4 y, BMI = 25.3). Blood was collected at 0, 0.5, 1, 1.5, 2, 2.5, 3, 4, 6, 8, 10, and 24 h after consumption of a muffin made with either 20 g AV‐enriched oat bran (containing 46.2 mg total AVs) or regular oat bran. After removal of lipids with hexane, AV phase II metabolites in the defatted plasma were extracted twice with ethyl acetate, dried under N2, and reconstituted in 55% methanol for analysis on an Agilent UPLC‐QToF‐MS equipped with an Agilent Zorbax Extend C‐18 RR HT column (2.1 × 50 mm, 1.8 μm). Utilizing the MassHunter Qualitative Analysis software, we identified with a confidence score ≥80 for aglycones AV‐O and ‐A, and ≥90 for methylated AV‐O. Their detection is consistent with plasma AV results obtained after the removal of the glucuronide and sulfate conjugates that AV‐O and ‐A were the most abundant isomers. AV‐A was detected at fewer time points than AV‐O probably due to its much lower circulating concentrations. AV‐A had a maximum concentration (Cmax) of 2.2 ± 1.6 ng/mL with the time to reach Cmax (Tmax) at 1.9 ± 0.8 h. The Cmax for AV‐O was 6.4 ± 3.9 ng/mL and the Tmax was 2.6 ± 0.8 h, as well as 14.0 ± 8.2 ng/mL AV‐O equivalents and 1.9 ± 0.7 h for methylated AV‐O. Though AVs are extensively transformed to phase II metabolites, our analyses reveal the presence of AV‐O and ‐A aglycones in the circulation. Ours is the first report of a methylated AV metabolite following acute AV consumption. Future studies are warranted to explore the potential bioactivity of methylated AVs.Support or Funding InformationSupported by USDA and PepsiCo

Gas-phase ions produced by freezing water or methanol for analysis using mass spectrometry.

Introducing water or methanol containing a low concentration of volatile or nonvolatile analyte into an inlet tube cooled with dry ice linking atmospheric pressure and the first vacuum stage of a mass spectrometer produces gas-phase ions even of small proteins that can be detected by mass spectrometry. Collision-induced dissociation experiments conducted in the first vacuum region of the mass spectrometer suggest analyte ions being protected by a solvent cage. The charges may be produced by processes similar to those proposed for charge separation under freezing conditions in thunderclouds. By this process, the surface of an ice pellet is charged positive and the interior negative so that removal of surface results in charge separation. A reversal of surface charge is expected for a heated droplet surface, and this is observed by heating rather than cooling the inlet tube. These observations are consistent with charged supercooled droplets or ice particles as intermediates in the production of analyte ions under freezing conditions.

Quantitation of tenofovir and emtricitabine in dried blood spots (DBS) with LC–MS/MS

A reversed-phase high performance liquid chromatographic (LC), tandem mass spectrometry (MS/MS) assay for the determination of tenofovir (TFV) and emtricitabine (FTC) in dried blood spots (DBS) from human whole blood was developed and validated. Whole blood samples were spotted, dried, and a 3mm punch was extracted with methanol for analysis by LC–MS/MS utilizing stable isotope labeled internal standards. The assay was validated over the range of 2.5–1000ng/mL for TFV and 2.5–5000ng/mL for FTC. The method was accurate (within ±15% of control) and precise (coefficient of variation ≤15%) for hematocrit concentrations ranging from 25% to 76%; using edge punches vs. center punches; and spot volumes of 10–50μL. Analytes were stable for five freeze/thaw cycles and up to 6 days at room temperature, whereas long-term storage required −20°C or −80°C. Comparison of TFV and FTC in DBS vs. plasma yielded r2≥0.96, indicating that DBS can be used as a plasma alternative for pharmacokinetic analyses in vivo.

Monoclonal Antibody-Based Immunoassay for the Detection of Maduramicin in Chicken Tissues

This paper reports on the preparation of a monoclonal antibody (Mab) against maduramicin (MD) and the development of a simple and sensitive ELISA for MD in chicken tissues. MD was conjugated to bovine serum albumin for immunogens and ovalbumin for coating antigens by mixed anhydride (MA) and active ester (AE) methods. Hybridoma cells were generated using spleen cells from a mouse immunized with MD-BSA conjugate. After screening with ELISA, the Mab with high anti-interference ability and high affinity was selected, and it exhibited negligible cross-reactivity with other usual-used polyether antibiotics. After optimization, the developed ELISA showed an IC50 value of 2.12 ± 0.46 ng/ml (n = 20). Chicken muscle and liver samples were extracted with methanol-8% NaCl solution (4:1) and then directly diluted with PBS-10% methanol for analysis. The recoveries of MD from spiked chicken tissues at levels of 40–480 µg/kg ranged from 81.3% to 91.3% with variation coefficients of 5.2–12.1%, and the detection limits were 6.5 µg/kg in muscle and 9.2 µg/kg in liver, respectively (n = 20).

A Caenorhabditis elegans Glycolipid-binding Galectin Functions in Host Defense against Bacterial Infection

Galectins are a family of beta-galactoside-binding proteins that are widely found among animal species and that regulate diverse biological phenomena. To study the biological function of glycolipid-binding galectins, we purified recombinant Caenorhabditis elegans galectins (LEC-1-11) and studied their binding to C. elegans glycolipids. We found that LEC-8 binds to glycolipids in C. elegans through carbohydrate recognition. It has been reported that Cry5B-producing Bacillus thuringiensis strains can infect C. elegans and that the C. elegans Cry5B receptor molecules are glycolipids. We found that Cry5B and LEC-8 bound to C. elegans glycolipid-coated plates in a dose-dependent manner and that Cry5B binding to glycolipids was inhibited by the addition of LEC-8. LEC-8 is usually expressed strongly in the pharyngeal-intestinal valve and intestinal-rectal valve and is expressed weakly in intestine. However, when C. elegans were fed Escherichia coli expressing Cry5B, intestinal LEC-8::EGFP protein levels increased markedly. In contrast, LEC-8::EGFP expression triggered by Cry5B was reduced in toxin-resistant C. elegans mutants, which had mutations in genes involved in biosynthesis of glycolipids. Moreover, the LEC-8-deficient mutant was more susceptible to Cry5B than wild-type worms. These results suggest that the glycolipid-binding lectin LEC-8 contributes to host defense against bacterial infection by competitive binding to target glycolipid molecules.

Lipidomic Analysis of Dynamic Eicosanoid Responses during the Induction and Resolution of Lyme Arthritis

Eicosanoids and other bioactive lipid mediators are indispensable regulators of biological processes, as demonstrated by the numerous inflammatory diseases resulting from their dysregulation, including cancer, hyperalgesia, atherosclerosis, and arthritis. Despite their importance, a robust strategy comparable with gene or protein array technology for comprehensively analyzing the eicosanoid metabolome has not been forthcoming. We have developed liquid chromatography-tandem mass spectrometry methodology that quantitatively and comprehensively analyzes the eicosanoid metabolome and utilized this approach to characterize eicosanoid production during experimental Lyme arthritis in mice infected with the bacterium Borrelia burgdorferi. Eicosanoids were extracted throughout infection from the joints of Lyme arthritis-resistant and -susceptible mice and subjected to lipidomic profiling. We identified temporal and quantitative differences between these mouse strains in the production of eicosanoids, which correlated with differences in arthritis development. The eicosanoid biosynthetic enzyme cyclooxygenase (COX)-2 has been implicated in the regulation of Lyme arthritis pathology, and subsequent lipidomic profiling of B. burgdorferi-infected COX-2(-/-) mice identified reductions not only in COX-2 products but, surprisingly, also significant off-target reductions in 5-lipoxygenase metabolites. Our results demonstrate the utility of a comprehensive lipidomic approach for identifying potential contributors to disease pathology and may facilitate the development of more precisely targeted treatment strategies.

Characterization and quantification of karlotoxins by liquid chromatography–mass spectrometry

Karlodinium veneficum is a cosmopolitan dinoflagellate with a worldwide distribution in mesohaline temperate waters. The toxins from K. veneficum, or karlotoxins (KmTxs), which have been implicated in fish kill events, have been purified from monoalgal cultures, and shown to possess hemolytic, cytotoxic and ichthyotoxic activities. Three karlotoxins (KmTx 1–1, KmTx 1–3 and KmTx 2) have been isolated from two different North American strains of K. veneficum and characterized using liquid chromatography–mass spectrometry (LC–MS). KmTx 1 karlotoxins have a UV absorption maximum ( λ max 225 nm) at lower wavelengths than KmTx 2 karlotoxins ( λ max 235 nm). The exact masses and predicted empirical formulae for the karlotoxins (KmTx 1–1, 1308.8210, C 67H 120O 24; KmTx 1–3, 1322.8637, and C 69H 126O 23; KmTx 2, 1344.7938, C 67H 121ClO 24) were determined using high resolution mass spectrometry. Although the individual toxins produce a single peak in reverse phase high performance liquid chromatography (HPLC), MS revealed congeners co-eluting within each peak. These congeners could be separated under normal phase chromatography and revealed a single hydroxylation being responsible for the mass differences. Multistage MS (MS n ) showed that the three KmTxs and their congeners share a large portion of their structures including an identical 907 amu core fragment. These data were used to develop a quantitative LC–MS assay for karlotoxins from cultures and environmental samples. The sensitivity afforded by MS detection compared to UV absorbance allowed toxin quantification at 0.2 ng when injected on column. Aqueous solutions of karlotoxins were found to quantitatively adsorb to PTFE and nylon membrane filters. Aliquots from whole cultures or environmental samples could be concentrated and desalted by adsorption to PTFE syringe filters and karlotoxins eluted with methanol for analysis by LC–MS. This simplified solid phase cleanup afforded new data indicating that each karlotoxin may also exist as sulfated derivatives and also provided a rapid detection method for karlotoxin from environmental samples and whole cultures.

Bioassay-based risk assessment of complex mixtures

In order to compare a standard chemical-based risk assessment with in vitro genotoxicity assays, two complex environmental mixtures from a wood-preserving site were analyzed in the Salmonella/microsome and E. coli prophage induction assays. Using GC/MS, sample 003 was found to contain relatively low levels of polycyclic aromatic hydrocarbons (PAHs) and elevated levels of polychlorinated dibenzo- p-dioxins (PCDDs), while sample 005 had higher levels of PAHs and relatively low levels of PCDDs. The complex mixtures were sequentially extracted with methylene chloride and methanol for analysis in Salmonella, or extracted with a 1:1 hexane:acetone mixture for analysis in the E. coli prophage induction assay. At a dose of 1.0 mg/plate in Salmonella strain TA98 with metabolic activation, the methanol extract of sample 003 induced 197 net revertants, while sample 005 induced 436 net revertants. In the prophage induction assay, with activation, the hexane:acetone extract of sample 003 induced a genotoxic response that was slightly lower than that observed with sample 005. The estimated incremental carcinogenic risk for ingestion of PAHs was 1.5E – 3 for sample 003, while for sample 005 the estimated risk was 1.5E – 2. Thus, the sample which induced the maximum response in both bioassays also had the highest estimated cancer risk. However, the frequency of PAH-DNA adducts in both skin and liver tissues was appreciably higher with sample 003 than with sample 005. A combined testing protocol, using both biological and chemical analysis, therefore provides more accurate information from which to assess risk than the use of either method alone.

Residue Level Determination of the Aquatic Herbicide Fluridone and a Potential Photoproduct (N-Methylformamide) in Water

Methods are presented for the determination of the aquatic herbicide fluridone and N-methylformamide (NMF), a potential photoproduct of fluridone which has thus far been observed only in a laboratory aqueous photolysis study. Water samples are passed through a Sep-Pak C18 cartridge to extract fluridone while NMF passes unretained through the cartridge with the water. Fluridone is eluted from the cartridge with methanol, which is then concentrated for analysis by liquid chromatography with UV detection at 313 nm. The water eluate containing NMF is concentrated for analysis by rotary vacuum evaporation at 40-46 degrees C. Methanol is added to help evaporate the water, and glycerol is added as a keeper solvent to retain NMF during the evaporation. The residual NMF is dissolved in methanol for analysis by gas chromatography with a Hall electrolytic conductivity detector operated in the nitrogen mode. The assay limits of detection are 1 and 2 ppb for fluridone and NMF, respectively. Recoveries averaged 98% for fluridone at 1-400 ppb and 87% for NMF at 2-50 ppb.

Determination of tibric acid and clofibrate in animal feed, wastewater and hunan urine.

Analytical chemical procedures are described to determine residues of the drugs clofibrate and tibric acid in animal feed, wastewater, and human urine. Clofibrate was extracted from animal feed and human urine with hexane, whereas residues from wastewater were collected on a Sep-PakTM then eluted with methanol for analysis. Clofibrate residues from the feed, wastewater, and urine were analyzed by high-pressure liquid chromatography (HPLC) with minimum detectable levels (MDL) of about 40, 0.5 and 1.0 ppb, respectively. Tibric acid was extracted from animal feed with 90% methanol and 10% 0.1 N NaOH, whereas wastewater and human urine were acidified with 12 N HCl and then extracted with benzene. The MDL for tibric acid in feed by electron capture/gas chromatography (EC/GC) and HPLC were about 40 ppb and 2.0 ppm, respectively. Residues from these extracts that contained more than 5 ppm of tibric acid were analyzed by HPLC, whereas GC was required for levels below 5 ppm. The GC procedures, which required that tibric acid be derivatized (methylated) prior to analysis, had MDL of 0.1 and 1.0 ppb for wastewater and human urine, respectively. Data are also presented concerning partition values, stability of the compounds in animal feed, and recoveries of the compounds from the three substrates.