Sample Preparation and Extraction

Sample extraction and preparation remain the most time-consuming and error-prone steps in the analytical process, but these are crucial procedures because food scientists need to isolate and concentrate a wide variety of analytes from complex and varied matrices. Advances in sample extraction and preparation in chemical analysis have only in the past several years been given critical consideration as an important component in obtaining reliable and robust analytical results. This situation is also true of analyses carried out in other industrial sectors (e.g. chemical and microbiological contaminants in food, agriculture and environment). 

Some of the factors mentioned earlier which introduce error when calibrating with external standards only, can be compensated for by introducing a constant amount of an internal standard in both the unknown and the standard calibration samples. This should be a compound with similar chemical nature to the analytes, so that it will pass through the sample extraction and preparation procedure similarly. In general it should elute in the chromatographic system close to the other peaks in the system, but separated from all of them, so it can be identified and measured accurately. One calculates the ratio of the peaks responses to their concentrations. Then the ratio for each analyte. A, peak is compared to ratio of the internal standard, IS peak, to give the relative response factor (RRF), for the analyte to... 

The optimization of MS/MS for amino acids in DBS was the first step in the development and validation of NBS [2]. The reason that an amino acids newborn screen was developed first rather than an AC screen was the requirement for validation of using a DBS for analysis and quantification by MS/MS. By choosing a compound that is MS friendly (i.e., positive ion, small molecule, previous MS analyses) and is also a key metabolite in NBS, it was more likely that validation of the assay would be achieved and that validation accepted. Phe is that MS-friendly metabolite used in the detection of PKU and screened for by all 50 US. states. It was likely that development of the Phe assay using MS/MS would be similar to that for ACs, since chemically speaking they are not that different in terms of molecular weight and the fact that they both have basic and acid groups. Sample extraction and preparation are also likely to be similar. So the initial attempts at an assay could mirror that used for plasma ACs. 

Protocols for mass spectrometric peptide analysis are common to modern and archaeological samples. The only significant difference in the analytical protocol used for archaeological samples is the need for careful sample extraction and preparation. In addition, the usefulness of archaeological proteomics requires the development of a database for archaeological peptide data, which will expand and increase over time but is currently relatively small. 

The procedure for proteomic study of archaeological proteins follows the general principles of conventional proteomics analysis, which can be divided into four main areas sample extraction and preparation, enzymatic digestion, LC separation, or sample plate spotting for... 

Assume a sample is delivered to the lab that is suspected to contain cocaine, lido-caine, benzocaine, and procaine. What would be the best method of sample extraction and preparation ... 

LC-GC, therefore, shows promise for forensic science applications, reducing sample handling and preparation steps by essentially using an on-line LC column in place of one or more extraction steps. This is followed by a traditional high resolution GC analysis. The methods described here for pesticides and hormones could be readily adapted to a variety of analyses, especially those involving fatty matrices. Such as tissues, food or blood. 

Another difference between determinative and confirmatory method trial procedures is the way in which sample extracts are prepared for analysis. Most current methods submitted for review use the same sample extraction technique for both the determinative and confirmatory procedures. In those cases where the same extraction technique is used, the sponsor may provide the prepared extract to the FDA laboratory for analysis. Any problems with the extraction procedure will have been corrected during the determinative method trial. 

Sample Preparation Liquid Samples, Extracts, and Solutions of Solids... 

Sample collection and preparation for the analysis of 1,2-dibromoethane in foods includes the purge-and-trap method, headspace gas analysis, liquid-liquid extraction, and steam distillation (Alleman et al. 1986 Anderson et al. 1985 Bielorai and Alumot 1965, 1966 Cairns et al. 1984 Clower et al. 1985 Pranoto-Soetardhi et al. 1986 Scudamore 1985). GC equipped with either ECD or HECD is the technique used for measuring 1,2-dibromoethane in foodstuffs at ppt levels (Clower et al. 1985 Entz and Hollifield 1982 Heikes and Hopper 1986 Page et al. 1987 Van Rillaer and Beernaert 1985). 

The tissue of interest should be removed from the animals using equipment that has been treated to ensure that it is RNase-free. The surface hair of the animals can be soaked in 70% ethanol to minimize the inclusion of hair or dander in the isolated tissues. A sterile disposable Petri plate, placed on top of a bed of crushed ice, is a suitable RNase-free surface for microdissection. Immediately after isolation from the animal, individual samples of tissue can be flash frozen by immersion in liquid nitrogen and stored in cryovials in liquid nitrogen or at -70°C until all samples from an experimental set have been obtained. RNA extraction and preparation of single-stranded cDNA can then be performed simultaneously on all samples. This will minimize differences between RNA populations that result from differences in sample preparation. 

Animals, usually rodents, are exposed to the test substance by an appropriate route, usually by gavage or by intraperitoneal injection. Each treated and control group must include at least five animals per sex. Positive controls should produce micronuclei in vivo at exposure levels expected to give a detectable increase over background. No standard treatment schedule (i.e., 1, 2, or more treatments at 24-h intervals) has been recommended. Three dose levels are generally used these should cover a range from the maximum to little or no toxicity. The erythrocytes are sampled from the bone marrow and/or peripheral blood of the animals. If bone marrow is used, the animals are sacrificed at appropriate times after treatment, the bone marrow extracted, and preparations made and stained. When peripheral blood is used, the blood is collected at appropriate times after treatment and smear preparations are made and stained. Preparations are analyzed for the presence of micronuclei. An increase in the frequency of micronucleated polychromatic erythrocytes in treated animals is an indication of induced chromosome damage. 

Filter if necessary and take a precise aliquot of the sample extract and dilute this until its concentration falls at approximately the mid-point of the calibration series prepared using the analytical standard. 

Analysis Is carried out on a cream stated to contain 2% w/w of both miconazole and hydrocortisone. An ODS column is used with a mobile phase consisting of acetonitrile/ acetate buffer pH 4.0 (70 30) and the eluent is monitored at 220 nm. A narrow range calibration curve, within 20% of the expected concentration of each analyte in the sample extract was prepared for each analyte by plotting the ratio of the areas of the analyte peaks against fixed amounts of the internal standards for both analytes. The internal standards used were econazole and hydrocortisone 21-acetate for miconazole and hydrocortisone, respectively. 

Sample extraction and hydrolysis details e.g., solvent extraction after freeze drying, with optimized acid or enzymatic hydrolysis Preparation of flavonoid standards and use of internal standards Chromatographic separation and detection method used, ideally RP-HPLC with UV or fluorescent detection Outline of quality assurance procedures employed... 

Analytical sample preparation Storage conditions prior to analysis Nature of sample analyzed Sample extraction and hydrolysis... 

Once samples are acquired, voucher specimens must be maintained according to standard accepted methods and the collected specimens must be extracted or otherwise processed to prepare samples for biological evaluation. The goal of sample handling and preparation is to select for positives (remove nuisance compounds), prepare the samples to be compatible with existing (and future) bioassays, and store both the collected unprocessed material and the processed samples in a manner that is easily retrievable and maximizes stability. 

Procedures for isolation and measurement of lipids in foods include exhaustive Soxhlet extraction with hexane or petroleum ether (AOAC, 1995 see Basic Protocol 1), chloro-form/methanol (Hanson and Olley, 1963 Ambrose, 1969), chloroform/methanol/water (Folch et al., 1957 Bligh and Dyer, 1959 see Basic Protocol 2 and Alternate Protocol 2), acid digestion followed by extraction (see Basic Protocol 4), or, for starchy material, extraction with n-propanol-water (e.g., Vasanthan and Hoover, 1992 see Basic Protocol 3). Each method has its own advantages and disadvantages and successful measurement of lipid content is often dictated by the type of sample and extraction medium employed. Commercial extraction and preparation of edible oils are explained in the literature (Williams, 1997). 

DESI Desorption electrospray ionization (DESI) is a recently developed technique that permits formation of gas-phase ions at atmospheric pressure without requiring prior sample extraction or preparation. A solvent is electrosprayed at the surface of a condensed-phase target substance. Volatilized ions containing the electrosprayed droplets and the surface composition of the target are formed from the surface and subjected to mass analysis (Takats et al., 2005 Wiseman et al., 2005 Kauppila et al., 2006). 

Preparation of standard, sample extraction and clean up Limit of detection (LOD), limit of quantitation (LOQ), and recovery (Rec) References... 

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