Acid-base extractions, sample fractionation

Solvent extraction of the sample is also frequently used in the analysis of particulate matter. Through the appropriate choice of solvents, the organics can be separated into acid, base, and neutral fractions, polar and nonpolar fractions, and so on. This grouping of compounds according to their chemical properties using extraction techniques simplifies the subsequent analysis. Each fraction can then be analyzed by GC-MS, with the GC retention time and the mass spectrum used for identification and measurement. 

One particular advantage of this extraction scheme is that large volumes of the sample can be processed on site with no sample manipulation. The sample is pumped through a 0.3 fim Balston microfiber filter tube (glass-fiber filter) directly onto the column array. This scheme fractionates the organic matter present in the water into hydrophobic and hydrophilic acid, base, and neutral fractions. Suspended sediment is also retained, and an extraction procedure for this material is presented. Organic matter from surface water... 

In our work at Leiden University, apart from HPLC-photodiode array (PDA) detection, HPLC-electrospray mass spectrometry was used as characterization and identification tools. A semipurified taxine extract obtained with acid/base extraction of T. baccata needles was analyzed in reversed phase nine taxines, one taxinine, and six taxanes were found present in the sample. Furthermore, 10-deacetylbaccatin 111 (paclitax-el s main precursor) and other taxanes were also found in the extract. Identification of the peaks was made with online liquid chromatography/mass spectrometry (LC/MS) and off-line nuclear magnetic resonance (NMR) following fraction collection at the end of the HPLC. Retention and spectral data of the identified peaks were used as tool to screen for taxines and taxi-nines in plant and cell culture extracts. Several... 

A technical challenge with this step is to achieve RNA extraction of uniform quality and efficiency for each fraction. This is because the amount of RNA in each sucrose gradient fraction varies considerably and the high concentration of sucrose in the bottom fractions interferes with phase separation in typical phenol-based extraction steps. To address these problems, we spike each fraction with an aliquot of a foreign (control) RNA, which can be used later to correct for differences in RNA recovery (and reverse transcription efficiency) between samples. We then remove sucrose from the samples by precipitation of total nucleic acid and protein with ethanol. To purify RNA, a standard Trizol (Invitrogen) extraction is performed as outlined later (also see product insert). 

Phenolic compounds have also been oxidatively polymerized to humic substances by clay minerals (29) and by the mineral fraction of a latasol (66). After a 10-day equilibration period, montmoril-lonite and illite clay minerals yielded 44 to 47% of the total added phenolic acids as humic substances whereas quartz gave only 9%. Samples of a latasol yielded over 63% of the total amount, from mixtures in varied proportion, of mono-, di- and trihydroxy phenolic compounds as humic substances (66). Extractions of the reaction products yielded humic, fulvic, and humin fractions that resembled soil natural fractions in color, in acid-base solubility, and in infrared absorption spectra. Wang and co-workers (67) further showed that the catalytic polymerization of catechol to humic substances was, enhanced by the presence of A1 oxide and increased with pH in the 5.0 to 7.0 range. Thus the normally very reactive products of Itgnin degradation can be linked into very stable humic acid polymers which will maintain a pool of potentially reactive phytotoxins in the soil. 

No sample fractionation procedure is given in the protocol, but several possible techniques were included in the literature review, including acid/base-neutral extraction, solvent fractionation (8), column chromatography, TLC, and HPLC (presented elsewhere in this chapter). 

The concentrated extract is then split. To one portion, 1.0 mL of DMSO is added, and the dichloromethane is removed under a stream of nitrogen. The resulting DMSO solution is used for the direct assay of the extract. If fractionation of the remaining extract is required, the investigator is given the option of using the acid/base-neutral extraction scheme described in step 3 of the nonaqueous liquid protocol or the HPLC technique described in the sample fractionation methods section. 

Niederer [100] used ion trap mass spectrometry and negative ion chemical ionisation to determine nitro- and oxypolyaromatic hydrocarbons in soils. Meyer et al. [101] have described a simple and reproducible method which provides the simultaneous determination of polycyclic aromatic hydrocarbons and het-eropolycyclic aromatic hydrocarbons (N, S, O) and their metabolites in contaminated soils. Contaminants extracted from the soil sample were separated by polarity and acid-base characteristics using solid-phase extraction on silica gel and a strong basic anion exchange material. A subfraction containing PANHs and neutral metabolites was subsequently fractionated into neutral and basic... 

In addition to the need to monitor known problematic compounds, newer compounds are being identified as potential threats to humans and as such need to be monitored in the atmosphere. For example, researchers reported (10) that several chemical and instrumental analyses of HPLC fractions provided evidence for the presence of /V-nitroso compounds in extracts of airborne particles in New York City. The levels of these compounds were found to be approximately equivalent to the total concentrations of polycyclic aromatic hydrocarbons in the air. Since 90% of the N-nitroso compounds that have been tested are carcinogens (10), the newly discovered but untested materials may represent a significant environmental hazard. The procedure involved collecting samples of breathable, particulate matter from the air in New York City. -These samples were extracted with dichloro-methane. Potential interferences were-removed by sequential extractions with 0.2 N NaOH (removal of acids, phenols, nitrates, and nitrites) and 0.2 N H2S04 (removal of amines and bases). The samples were then subjected to a fractional distillation and other treatments. Readers interested in the total details should consult the original article (10). Both thin-layer chromatography (TLC) and HPLC were used to separate the compounds present in the methanolic extract. 

The complexity of flavour isolates and vast differences in the relative concentrations of individual compounds make it difficult to identify trace components in such mixtures even with the use of efficient capillary columns. A common approach to simphfy the complexity of flavour isolates is to fractionate the sample into non-polar, moderately polar and strongly polar components by silica gel chromatography or separate the components into acidic, basic and neutral fractions by acid and base extractions. 

An important issue emerged from the results of experiments with 14C-labeled pyrene added to a pristine forest soil (Guthrie and Pfaender 1998) (1) extensive mineralization took place only in samples amended with a pyrene-degrading microbial community, (2) there was a substantially greater nonextractable fraction of label in soils containing either the natural or introduced microflora compared with an azide-treated control, (3) metabolites that could be released by acid and base extraction remained in the soil after 270 days of incubation. 

LIE can be performed simply using separatory funnels. The partition coefficient should therefore be large because ffiere is a practical Umit to the phase-volume ratio and the number of extractions. When the partition coefficient is small and the sample very dilute, a large volume must be handled and continuous liquid-liquid extractors should be used. The extractions then take several hours. Such extractors have been described in the literature [ 187]. The partition coefficient may be increased by adjusting the pH to prevent ionization of acids or bases or by forming ion pairs or hydrophobic complexes with metal ions, for example. The solubihty of analytes in the aqueous phase can be reduced by adding salts. Fractionation of samples into acidic, basic and neutral fractions can be attained by successive extractions at different pH [ 1 ]. 

Method 625 for Semivolatiles. This method is a solvent extraction method intended to determine as many of the organic semivolatile priority pollutants as possible. To accomplish this, the sample is serially extracted, first at a pH greater than 11 and then at pH 2. Figure 1 shows a flow diagram of the procedure. The two fractions, base-neutrals and acids, are independently determined by using two separate GC columns. The base-neutrals are determined on a 1.8-m X 2-mm i.d. glass column packed with Supelcoport (100-120 mesh) coated with 3%... 

Jince the time of Berzelius, chemists have proposed structures for the amorphous, black substance known as humic acid. In the past 150 years, much experimental work has appeared on the nature of humic acid, most of it based on classical chemical and microbiological studies. Very little information about the molecular structure of humic add has resulted from these studies however. Some of the problems plaguing investigators in this field have been (a) variation in the source of humic acid, (b) variation in the definition of humic fractions of soil and coal, (c) lack of crystallinity of the samples, (d) uncertainty of molecular weight measurements, (e) variation in extraction techniques, and (f) variation in elemental composition. The little unambiguous information that exists today is based on extensive degradation of the humic acid polymer and represents only a small fraction of the total molecule. 

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