Bioautography

Visualisation of the chromatograms has been reported using both chemical and microbiological means. Chemical spray reagents which have been described are listed in Table 16. The various organisms that have been employed for bioautography are given in Table 11. 

The use of bioautography for the quantitation of chromatograms has been described by many workers. Emilianowicz-Czerska and Herman228 separated neamine from neomycins B and C then quantitatively determined total neomycin B and C by bioautography with B.iubtZZZi. A linear response for concentrations between 0.25 and lOyg of neomycin was obtained. With a similar technique but using a mathematical analysis of the bioauto-... 

High voltage electrophoresis (HVE) in agar gel with detection by bioautography has been used with considerable success in some laboratories for identification of residues (1-6). This procedure has the advantage that all antibiotic substances detectable by bioautography can be classified on the basis of electrophoretic mobility. Further testing may be required for quantification and to... 

Tolerances for these compounds are generally O-.Ol ppm except for penicillin G in cattle (.05 ppm) and cephapirin in edible tissue (0.1 ppm) and milk (.02 ppm) (70). Many chromatographic methods have been described for determination of these compounds in clinical applications, but these methods are not sufficiently sensitive for residue analysis. The summary of methods in Table II includes one GLC (71), five TLC (72-76), and five HPLC methods (77-81). Four of the TLC methods use detection by bioautography. Three HPLC methods have been described for milk (77-79) and two for tissue (80,81). The HPLC methods described by Moats C7 ,80) and by Munns et al (77) are satisfactory for any penicillin with a neutral side-chain and this may be true with the procedure of Terada, et al. (81). The procedure of Terada and Sakabe (79) is also satisfactory for the aminopenicillin, amplclllin. The method of Munns et al (77) can also be used to detect the corresponding penicilloic acid metabolites. 

Penicillins form several major metabolites which are excreted in the urine (83,84). These metabolites are usually inactive microbio-logically and they would not be detected by the usual microbiological tests. There are no analytical methods for these metabolites in tissues and, therefore, little is known as to their occurrence and persistence in tissues. There are no methods available for identifying residues of some commonly used B-lactam antibiotics including carbenicillin and ticarcillin. For cephapirin and ampicillin, except for one HPLC method for ampicillin in milk (79) only TLC procedures (72-74,76) with detection by bioautography are reported. 

Weiss and MacDonald (87) recently reviewed methods for determination of ionophore antibiotics. lonophores approved for use in animal agriculture in the U.S. are lasalocid, monensin, and salinomycin. An HPLC ( ) and GLC-MS ( ) procedure have been described for lasalocid. For other ionophores, TLC-bioautography is the preferred procedure because of lack of any useful UV absorbance. However, a few TLC colorimetric procedures have been described for monensin residues in tissues (90-92). 

TLC-bioautography has been used in Canada since 1984 for the confirmation of tetracycline-positive in plant tests (65). However, TLC-bioautography is not quantitative and only gives direction to the analyst as to what confirmatory method of analysis should be used. Therefore, presumptive positives must be confirmed by physicochemical techniques that have been validated in terms of detection limit, precision, and accuracy. 

Determination of novobiocin in animal tissues and milk has been reported using either thin-layer chromatography-bioautography (14) or liquid chromatog-... 

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