Tissue analytical procedure

The therapeutically active dmg can be extracted from plant or animal tissue, or be a product of fermentation (qv), as in the case of antibiotics. Frequentiy, it is synthesized and designed to correlate stmcture with therapeutic activity. Pharmacologic activity is first tested on laboratory animals. When the results ate encouraging, physical and chemical properties are determined in the so-called preformulation stage, and analytical procedures are developed for quahty control (see Qualityassurance/qualitycontrol). 

This means that if all of the TCDD were retained, the level of TCDD would be less than 1 part per billion (ppb) in the whole animal. The lowest reported limit of detection for TCDD in whole tissue is 50 ppb (6). Thus, a guinea pig could be killed with TCDD, and it would be impossible to establish this fact with the analytical procedures in current use. 

By utilizing the HPLC method, it is possible to determine the level of each individual toxin in sample solutions. This provides a "toxin profile" that can be very useful in PSP toxin research studies. The ability to examine relative changes in toxin concentration and profile has greatly facilitated studies relating to toxin production by dinoflagellates, metabolism of toxins in shellfish, and movement of toxins up the food chain. Since the HPLC method is easily automated and requires only very small sample sizes (< 1 g tissue), it has clear advantages over other analytical procedures for the toxins in many research situations. Two examples of the utilization of HPLC for the study of the PSP toxins follow. 

The development of specific and reliable analytical procedures for the detection, location, and quantification of mineral particles in biological tissues (Henderson and Barr, 1988) has provided both the experimental techniques and additional evidence for detecting aluminosilicates in Alzheimer brains (Singhrao et al., 1990). The association of asbestos-related disease with severe... 

Because estrogenic mycotoxins usually occur at microgram per kilogram (pg/kg) levels there is special interest in analytical procedures for reliable detection of zearalenone and its metabolites between 10 and 100 pg/kg. In response to the risk of a great economic loss to the industry and the threat to human health as a result of exposure to zearalenone, several methods have been developed for the quantification of zearalenone and its metabolites in different foods, feeds, animal tissues, blood and urine. Detailed reviews have been given by Steyn et al. 1991 Betina 1993 Frisvad and Thrane 1993 Scott 1993 Steyn 1995 and Lawrence and Scott 2000. The determination of zearalenone in cereals can be divided into five steps grinding of the sample, extraction of the sample, clean-up, separation and detection. 

Monooxygenase Assays. Incubation media contained the following (final concentrations) 0.05M phosphate buffer, pH 7.A, glucose-6-phosphate (G-6-P, 2.3 mM), G-6-P dehydrogenase (3 units), NADP (0.23 mM), and KC1 (2.8 mM), and various tissue preparations. Substrates were added in small volumes (25 yl or less) of MeOH. Samples (1.1 ml) were shaken in a thermostated (usually at 22°C) water bath and reactions terminated by enzyme denaturation. Specific analytical procedures for aldrin epoxi-dation (13), 1 CH30-p-nitroanisole 0-demethylation (1A), and 3H-benzo(a)pyrene oxidation (15) have been described. 

In vivo oxidation activity may not be expressed in vitro due to inclusion of excessive amounts of "inactive" tissue in the various cell-free preparations. The resulting tissue dilution artifact renders activity unmeasurable due to the sensitivity of the analytical procedures. This consideration warrants further experimental evaluation. 

Liere P, Akwa Y, Engerer SW, Eychenne B, Pianos A, et al. 2000. Validation of an analytical procedure to measure trace amounts of neurosteroids in brain tissue by gas chromatography-mass spectrometry. J Chromatog B 739 301-312. 

Until the early 1970s analytical chemists could detect DBS residues in beef tissue, specifically liver, at a level of 5-10 parts DBS per one billion parts of beef (5-10 ppb). If DBS were present above this level it could be detected with existing analytical procedures, but it could not be found if it were present at any concentration from zero to 5 ppb. Under conditions of cattle dosing approved by the BDA, no residue of the drug could be found in the late 1960s. The drug could safely and legally be used. 

Rushing and Hansen (504) described a sensitive analytical procedure for screening and confirmation of residues of malachite green, gentian violet, and their leuco analogs in catfish and trout tissues using liquid chromatographic separation and electrochemical, diode array, and fluorescence detection in series. 

To purify a protein, it is essential to have a way of detecting and quantifying that protein in the presence of many other proteins at each stage of the procedure. Often, purification must proceed in the absence of any information about the size and physical properties of the protein or about the fraction of the total protein mass it represents in the extract. For proteins that are enzymes, the amount in a given solution or tissue extract can be measured, or assayed, in terms of the catalytic effect the enzyme produces—that is, the increase in the rate at which its substrate is converted to reaction products when the enzyme is present. For this purpose one must know (1) the overall equation of the reaction catalyzed, (2) an analytical procedure for determining the disappearance of the substrate or the appearance of a reaction product, (3) whether the enzyme requires cofactors such as metal ions or coenzymes, (4) the dependence of the enzyme activity on substrate concentration, (5) the optimum pH, and (6) a temperature zone in which the enzyme is stable and has high activity. Enzymes are usually assayed at their optimum pH and at some convenient temperature within the range... 

Animals taken from experimental or control groups for the analytical procedures were decapitated by guillotine. The brain of each animal was then removed as rapidly as possible, the total elapsed time seldom exceeding 20 seconds. Further processing of the tissue depended upon the constituent to be measured. 

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