Biological samples, extraction

Recently, most of the methods which have been used for the analysis of valproic acid in plasma, serum, cerebral spinal fluid, saliva, breast milk, and urine involve acidification of the biological sample, extraction into an organic solvent, and direct injection onto a gas-liquid chromatographic column (28, 29, 16, 30, 31, 32,... 

The sample was submitted to MAE using acetone-n-hexane (1 1, v/v) mixture for sediment samples and methanol 112 or methanolic 1 M KOH for biological samples. Extract solution was centrifuged and evaporated just to dryness. Residues were dissolved in 1 ml of n-hexane. Cleanup was performed on a Si02/Al203. PCBs were eluted with n-hexane... 

Pizarro, I. G6mez, M. Camara, C. Palacios, M.A. Arsenic speciation in environmental and biological samples Extraction and stability studies. Anal. Chim. Acta 2003, 495(1), 85. 

Another possible mechanism involves increases in the surface tension of the droplets as a result of high concentrations of co-eluting compounds (Mallet 2004). This effect would reduce the rate of solvent evaporation, and thus also the probability that the droplet will reach a sufficiently small size that ion evaporation can occur, and also increase the difficulty of ion evaporation itself via the term in the rate constant, see Equations [5.6-5.7]. Another effect that seems to be particularly important for biological sample extracts arises from the presence of involatile solutes, believed (King 2000) to cause ionization suppression not only via a strong effect on surface tension but also via co-precipitation of analyte as the droplets shrink in size. 

In the case of water or liquid biological samples, extraction is the next step after filtration, while in case of solid samples it comes after homogenization. Many different devices have been used for extraction procedures. The most important techniques in this field are the classical methods of extraction, liquid-liquid extraction, solid phase extraction and liquid chromatographic techniques. The extraction of solid and liquid samples of environmental and biological origin is described separately. 

If the lipids of interest are minor constituents of the sample or present in relatively low concentrations in a complex biological sample, extraction, isolation, and concentration steps usually precede TLC. Numerous sample preparation methods are available, and personal choice often dictates which one will be used. Of all the procedures, that of Folch et al. (70), ora modification of the original procedure, is used more frequently than any other. Table 4 lists numerous procedures used for the sample preparation of lipids. 

Owing to the light and air sensitivity of the carotenoids and retinoids, sample handling is a critical issue. It is recommended to conduct extraction of these materials with peroxide-free solvents, to store biological samples at —70° C under argon and in the dark, to perform the analysis under yellow light, and to use reference compounds of high purity (57). 

Presently, the on-line coupling of NPLC and GC via heart-cutting is an established procedure which has been used successfully for several bioanalytical applications. Obviously, dfrect analysis of aqueous samples is not possible by NPLC, and therefore, a solvent switch by a sample pretreatment step (e.g. liquid-liquid extraction or SPE) is always requfred when biological samples are analysed by NPLC-GC. 

Zebiihr et al. (29) developed an automated system for determining PAHs, PCBs and PCDD/Fs by using an aminopropyl silica column coupled to a porous graphitic carbon column. This method gives five fractions, i.e. aliphatic and monoaromatic hydrocarbons, polycyclic aromatic hydrocarbons, PCBs with two or more ortho-chlorines, mono-ort/io PCBs, and non-ortho PCBs and PCDD/Fs. This method employed five switching valves and was successfully used with extracts of sediments, biological samples and electrostatic filter precipitates. 

Reliable analytical methods are available for determination of many volatile nitrosamines at concentrations of 0.1 to 10 ppb in a variety of environmental and biological samples. Most methods employ distillation, extraction, an optional cleanup step, concentration, and final separation by gas chromatography (GC). Use of the highly specific Thermal Energy Analyzer (TEA) as a GC detector affords simplification of sample handling and cleanup without sacrifice of selectivity or sensitivity. Mass spectrometry (MS) is usually employed to confirm the identity of nitrosamines. Utilization of the mass spectrometer s capability to provide quantitative data affords additional confirmatory evidence and quantitative confirmation should be a required criterion of environmental sample analysis. Artifactual formation of nitrosamines continues to be a problem, especially at low levels (0.1 to 1 ppb), and precautions must be taken, such as addition of sulfamic acid or other nitrosation inhibitors. The efficacy of measures for prevention of artifactual nitrosamine formation should be evaluated in each type of sample examined. 

In the past decade, new sample extraction techniques have been introduced to meet stricter criteria in the areas of food and agriculture, for example, enviromnentally friendly, non-toxic, fast, automated, robust, and cost-efficient techniques. Accelerated solvent extraction (ASE) and pressurized liquid extraction (PEE) are two methods developed for the extraction of chemicals of interest " and provide high yields and efficiency from a wide range of botanical, - animal, and biological samples. ASE and PLE combine solvents at elevated temperatures (40 to 200°C)... 

The use of ethyl acetate was suggested by Oszmianski and Lee (1990) to wash out phenolics other than anthocyanins. Finally, a relatively pure anthocyanin extract can be removed from the colnmn with acidified methanol (0.1% HCl). Anthocyanin extracts can be enriched in this way by use of solid phase purification, which is especially helpful for diluted samples such as biological samples. Two factors in the nse of these purification techniques are the stability of anthocyanins to the conditions nsed and the ease of anthocyanin recovery from the column. ... 

Ham GJ, Stradling GN, Breadmore SE. 1977. Determination of americium and curium in biological samples of extraction and liquid scintillation. Anal Chem 49 1268-1270. 

This chapter on analysis of methylxanthines is divided into three sections historical methods, current analytical methods for foods, and current methods for biological samples which can include plasma, blood, urine, cell extracts, and other potential samples of biological significance. 

Acrylonitrile metabolites have been measured in blood and urine, but, except for measurement of thiocyanate, these methods have not been developed for routine monitoring of exposed humans. Supercritical fluid extraction/chromatography and immunoassay analysis are two areas of intense current activity from which substantial advances in the determination of acrylonitrile and its metabolites in biological samples can be anticipated. The two techniques are complementary because supercritical fluid extraction is especially promising for the removal of analytes from sample material and immunoassay is very analyte-selective and sensitive (Vanderlaan et al. 1988). 

Sperling [133] has reported extensively on the determination of cadmium in seawater, as well as in other biological samples and materials. He added ammonium persulfate, which permitted charring seawater at 430 °C without loss of cadmium. For workbelow 2 pg/1 cadmium in seawater he recommended extraction of the cadmium to separate it from the matrix [126,134,135]. He found no change in the measured levels over many months when the seawater was stored in high-density polyethylene or polypropylene. 

Han, X. and Gross, R. W. Global analyses of cellular lip-idomes directly from crude extracts of biological samples by ESI mass spectrometry a bridge to lipidomics. /. Lipid Res. 44 1071-1079, 2003. 

PCBs in biological samples are usually extracted by a Soxhlet column and with a nonpolar solvent such as hexane. The sample is first mixed with sodium sulfate to remove moisture. The extraction of PCBs from sediments was tested with sonication, with two sonications interspersed at a 24-h quiescent interval, with steam distillation, or with Soxhlet extraction (Dunnivant and Elzerman 1988). Comparison of the recoveries of various PCB mixtures from dry and wet sediments by the four techniques and the extraction efficiency of four solvents showed that the best overall recoveries were obtained by Soxhlet extraction and the two sonication procedures. In comparisons of solvent systems of acetone, acetonitrile, acetone-hexane (1+1), and water-acetone-isooctane (5+1.5+1), recoveries of lower chlorinated congeners (dichloro- to tetrachloro-) were usually higher with acetonitrile and recoveries of higher chlorinated congeners (tetrachloro- to heptachloro-) extracted with acetone were superior (Dunnivant and Elzerman 1988). The completeness of extraction from a sample matrix does not seem to discriminate against specific isomers however, discrimination in the cleanup and fractionation process may occur and must be tested (Duinker et al. 1988b). 

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