Tetracyclines spectrophotometric methods

Direct spectrophotometric methods using fluorometric detection have been reported for screening of tetracycline in milk extracts (126,282), and three tetracyclines in tissues extracts (302, 310). In a different approach, Salinas et al. (212) described a fourth-derivative spectrophotometric method for the determination of oxytetracycline in honey. Although useful, these methods cannot differentiate among individual tetracycline antibiotics and are less sensitive than the screening tests used for regulatory purposes. 

Table XVII. Recovery of Tetracycline Antibiotics in Various Dosage Forms by the Nonaqueous, Microbiological, and Spectrophotometric Methods... 

A microbiological assay is the method of choice (see p. 813) but spectro-photometric methods are also applicable. One spectrophotometric method is based on the formation of a yellow colour with an absorption maximum at 380 m//, when tetracycline is dissolved in 0-25N sodium hydroxide, another on the orange-brown colour (maximum absorption 490 m//) formed on mixing a dilute hydrochloric acid solution of the sample with ferric chloride solution. The former method which is described below is also applicable to oxytetracycline but not to chlortetracycline while the ferric chloride reaction, which is given in detail under oxytetracycline, applies to all three. 

The tetracycline which may be present is then converted to an anhydro derivative by heating in acid and is measured spectrophotometrically. Feldman et al. (84) developed the alkaline degradation method to measure CTC in fermentation mash and Spock and Katz (85) used this method to determine CTC in animal feed premixes. 

Following sample extraction and cleanup, tetracycline antibiotics may be separated by thin-layer or liquid chromatographic procedures, and quantified by spectrophotometric, fluorometric, or mass spectrometric detection systems. Direct spectrophotometric and capillary electrophoretic methods have also been described (Table 29.8). 

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