Bound residue determination

The chelate effect in proteins is also important, since the three-dimensional (3-D) structure of the protein can impose particular coordination geometry on the metal ion. This determines the ligands available for coordination, their stereochemistry and the local environment, through local hydrophobicity/hydrophilicity, hydrogen bonding by nearby residues with bound and non-bound residues in the metal ion s coordination sphere, etc. A good example is illustrated by the Zn2+-binding site of Cu/Zn superoxide dismutase, which has an affinity for Zn2+, such that the non-metallated protein can extract Zn2+ from solution into the site and can displace Cu2+ from the Zn2+ site when the di-Cu2+ protein is treated with excess Zn2+. 

It is questionable whether or not the value obtained by bioassays or the sequestration value can be used to define contaminant-bound residues. Ageing-sequestration relationships in the subsurface, as determined through bioavailability, may provide an... 

Applying Equation 8, the calculated 18 2/18 3 ratio was very similar to the experimentally determined ratio for all compounds except 2-(4-hydroxyphenyl)-4-chloro-5-dimethylamino-pyridazin-3-one (4.18 calculated vs. 0.84 experimental. Table III). This large difference could be explained by the well known ability of chemicals with phenolic HO- groups to form conjugates which result in bound residues. Therefore, repeating the process of calculation as shown in Equation 8, excluding the 4-chloro-5-dimethylamino-pyridazin-3-one with the 4-hydroxyphenyl substituent in two-position of the heterocycle, results in Equation 9 ... 

This nonextractable radioactivity was probably the result of covalent binding of the furazolidone intermediates to endogenous macromolecules. The bioavailability of these bound tissue residues from the above pig residue depletion study was determined by feeding rats lyophilized samples of liver and muscle tissues from animals sacrificed at 0 and 45 days after the last treatment (132). Results showed that the fraction of the bound residues bioavailable to rats was in the range 16-41%. The toxicological impact of these bioavailable bound residues has not been yet determined. 

Chen, R.F. 1968. DANSYL labeled proteins determination of extinction coefficient and number of bound residues with radioactive dansyl chloride. Anal. Biochem. 25, 412 116. 

Other assumptions and variables may also be involved in determining MRLs, including safety factors used in establishing ADIs, withdrawal times, the contribution of bound residues, and the bioavailability of residues. 

Several xenobiotics that are metabolized by GSH conjugation In plants, Including atrazine, propachlor, and PCNB, produce significant levels of bound residue (15). It appears that the bound residue may be formed from the GSH pathway with either a cysteine conjugate or a thiol as a precursor 1 5). The chemical nature of these bound residues has not been determined. 

Nearly all current methods of isolation and purification of lectins rely on affinity chromatography. Naturally, the characteristic ligand must be determined in advance. The properties of lectins can be used to precipitate macromolecules and to agglutinate some types of cells, be they plant or animal. The driving force of this reaction is the association with certain bound residues, generally monosaccharides, from the macromolecule or the cellular periphery. When this type of reaction is observed, the problem is to find the sugar that can inhibit activity at the lowest possible molar concentration. As in the case of immunochemical precipitations, this inhibition is due to the occupation of the recognition site by the small soluble molecule. 

Conditions see Table II Assay Procedures The activity of acid invertase (free and bound) was determined in plant material that was frozen in liquid nitrogen, freeze-dried, pulverized, homogenized with mercaptoethanol, centrifuged at 25,000g for 30 min (4WC) solution and residue were dialyzed (48 h, 4WC) and then incubated with sucrose (pH 4.7, 1 h, 30 0 and glucose was analyzed electrochemically. 

The results of these experiments in both aquatic systems and terrestrial systems may profitably be viewed against the extensive evidence for the persistence of agrochemicals in the terrestrial environment. Considerable effort has been directed to the issue of bound residues of agrochemicals (Calderbank 1989) and to its significance in determining both their biological effects and their persistence this is now fully accepted in contemporary thinking. At the... 

All sediment samples were pre-extracted with methanol, butanol and solutions of hexane/acetone. The principal analytical flow scheme applied to the samples for the detection and determination of bound residues is given in Fig. 2. 

Fig. 2 Flow scheme of the analytical procedure for determining bound residues...

Northcott GL, Jones KC (2000) Experimental approaches and analytical techniques for determining organic compounds bound residues in soil and sediment. Environ Poll 108, 19-43. 

Figure 4.3 Sandwich-immunoassay for the determination of bound pesticide residue. Schematic view. Antibody 1 (Ab 392) is immobilized on a polystyrene surface. Incubation of the analyte (bound residue). Addition of antibody 2 (Ab K1F4) detection by use of a third enzyme-labelled antibody 3. (From Ulrich, M.G.,. Anal. Chem., 354, 352, 1996. With permission.)... 

In the present study, three groups of six cattle (a total of eighteen) were administered the Captec device intra-ruminally via a fistula. Drug release rates were determined by HPLC assay of ABZ and its major metabolites in plasma as well as by physical measurement of the plunger movement. ABZ marker residue levels were determined using the established regulatory procedure in total liver, an ethyl acetate extractable fraction as well as the remaining intractable (bound) residue on each of three cattle at 0- and 5-Day withdrawal. 

A Soxhlet technique was used for determination of soil-bound DDT residues which may be released gradually from the soil increasing the insecticide load of the soil. The bound residues so released may be available for uptake by the biota. In this application, 50 g samples of air-dried soil, in triplicate, were extracted by three volumes of methanol in Soxhlet apparatus for 24 h (72 cycles). Sulphuric acid was used for clean-up of the extract since it did not affect DDT and its metabolites. The released residues were taken up in HPLC-grade methanol for analyses by HPLC. 

Immunochemical methods have also been applied to the detection of bound pesticide residues in soil. These are formed by binding of pesticides to the organic matter of the soil, mainly humic and fulvic acids, and cannot be analyzed using common extraction and assay methods. Hahn et al. used Fab fragments labeled with a fluorescent dye to detect nonextractable residues of atrazine in soil from corn fields. The fluorescence signal obtained was related to the amount of bound atrazine in native soil samples determined by GC after supercritical methanol extraction. A noncompetitive sandwich lA for the analysis of bound residues based on HA-Ab and triazine Ab was developed by Ulrich et HA was extracted from soil, bound to the plates by the HA-Ab and the nonextractable triazine residues were detected by... 

P. Ulrich, R. Niessner, Development of a Sandwich Immunoassay for the Determination of Bound Residues , Fresenius J. Anal. Chem., 354, 352-358 (1996). 

Bound residues of tetracyclines may occur in bones of slaughtered animals for months after treatment. Theoretically, these could reach the food chain via contaminated (mechanically deboned) meat or meat and bonemeal. The accumulation of tetracyclines in tissues is illustrated by the findings of Toutain and Raynaud for oxytetracycline in calves (Table 2.8). Concentrations of oxytetracycline were relatively high in liver and kidney compared to the extrapolated zero-time concentration for serum (4.2 mg/1). The time required for residues to deplete to 0.1 mg/1 in serum was 143 hr, considerably shorter than the time required for residues to deplete to 0.1 mg/kg in liver and kidney, but similar to the depletion time for muscle. The data nicely illustrate the importance of tissue elimination half-life in determining decrease to the 0.1 mg/kg concentration despite an almost three-fold higher initial concentration... 

Conneely A, Nugent A, O Keeffe M, Mulder PPJ, van Rhijn JA, Kovacsics L, Fodor A, McCracken RJ, Kenned DG, Isolation of bound residues of nitrofuran drugs from tissue by solid-phase extraction with determination by liquid chromatography with UV and tandem mass spectrometric detection, Ana/. Chim. Acta 2003 483 91-98. 

Bound residues were incubated at pH 1, 5, and 9 at a temperatrue of 37 °C and the extent of hydrolysis/solubilization was determined by quantification of the radioactivity released. An additional 3-9% TRR was solubilized from the (XVI) bound residue, but at pH 9, ca. 18% TRR was solubilized from the (XVII) bound residue. This may be due in part to the tendency of hemicellulose to be solubilized in basic conditions. Results are displayed graphically in Figure 4. 

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