Concerns about cross-allergenicity between sulfonamide antibiotics and nonantibiotic, sulfonamide-containing drugs persist and can complicate patients' drug therapy unnecessarily. No interaction between the human immune system and the sulfonamide functional group has been demonstrated. The immunologic determinant of type I, immediate hypersensitivity responses to sulfonamide antibiotics is the N1 heterocyclic ring. Nonantibiotic sulfonamides do not contain this structural feature. Non-type I hypersensitivity responses to sulfonamide antibiotics are largely attributable to reactive metabolites that may cause either direct cytotoxicity or immunologic response. Formation of these metabolites is a stereospecific process that occurs at the N4 amino nitrogen of the sulfonamide antibiotics, a structure also not found on any nonantibiotic sulfonamide drugs. The stereospecificity of these reactions implies that cross-reactivity with nonantibiotic sulfonamide-containing drugs is highly unlikely; this assertion is supported by recent literature. However, T-cell recognition of unmetabolized, nonhaptenated parent sulfonamide antibiotic appears to occur in a small subset of hypersensitive patients. Several of the severe cutaneous reactions associated with sulfonamide antibiotics are mediated by T cells. It is not known whether T-cell recognition of antibiotic is related to the sulfonamide functional group. Until the mechanism of this recognition is elucidated, cross-reactivity with nonantibiotic sulfonamides appears to remain at least theoretically possible.

A method for direct detection of Listeria monocytogenes in 25 ml of raw milk is presented. The detection limit can be situated between 10 and 5 CFU. The detection method is based on chemical extraction of the milk components and PCR amplification with two nested pairs of primers specific for Listeria monocytogenes. Listeria monocytogenes is a gram-positive pathogenic bacterium. Human listeriosis is frequently transmitted by the consumption of dairy products. The classical detection method for L. monocytogenes in milk and milk products involves enrichment for 48 h and subsequent colony formation on selective agar medium for 48 h, followed by a set of biochemical and morphological confirmation tests which typically take days to complete (8). In vitro amplification of specific DNA sequences by PCR allows direct detection and identification of the pathogen. Different PCR assays using primer sets derived from the listeriolysin O gene (3, 16), the Dth18 gene (18), and the iap gene (4) have been reported for L. monocytogenes. With PCRbased detection systems, even a single bacterium may be detected, as was reported for sediment (15) and water (1) samples. When DNA is extracted from complex food matrices like dairy products, the PCR may be 10 to 10 7 times less sensitive, depending on the type of product tested (7, 18). Sensitive detection in dairy products becomes possible when the bacterial cells are efficiently concentrated and purified from the food components prior to lysis. One way of achieving this is the magnetic immuno-PCR assay, which uses magnetic beads coated with specific monoclonal antibodies to concentrate Listeria cells out of the enrichment culture (6). The analysis time required to achieve a detection limit of 1 CFU of L. monocytogenes per g of cheese was approximately 55 h. Filtration was applied as an alternative method to concentrate L. monocytogenes cells from milk components prior to PCR identification (14). The filter was solubilized in chloroform, and the DNA was purified by phenol-chloroform extraction. No detailed information about the fat content, the somatic cell number, and the total background flora, which define the filtration properties of the milk analyzed, was given (5). We developed a direct detection method for L. monocytogenes in raw milk on the basis of chemical extraction of the milk components. This method supplies very pure DNA that is suitable for PCR. The sensitivity of detection is enhanced by application of a two-step PCR amplification procedure with two nested pairs of primers specific for L. monocytogenes. To raw milk (per milliliter), 200 l of NH 3 , 200 l of ethanol, 300 l of diethylether, 150 l of petroleum ether, and 0.07% sodium dodecyl sulfate (SDS) were added. Because of the toxicity of the reagents applied, working in a fume cupboard with protective gloves is recommended. The bacterial

Retention volumes for a group of sulfonamides obtained on three different HPLC columns were correlated with log P, pKa, and biological activity. Depending on the chromatographic system, good correlations can be obtained with log P or with biological activity, but not necessarily both, using the same chromatographic data.

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A direct detection method for Clostridium tyrobutyricum spores in up to 100 ml of raw milk is presented. The bacterial spores are concentrated by centrifugation after chemical extraction of the milk components. The vegetative cells are selectively lysed, and their DNA is digested and washed away. Afterwards, the DNA is liberated from the spores by microwave treatment. For the identification of the C. tyrobutyricum DNA, a two-step PCR method with two nested pairs of primers is used. The primers were derived from the 16S-23S rRNA spacer region of C. tyrobutyricum, and the specificity of each of them for C. tyrobutyricum is demonstrated. The detection limit can be estimated to be between 3 and 30 spores in 100 ml of raw milk.