Antibiotics, detection tetracyclines

Another subclass of proteases attacks internal peptide bonds and Hberates large peptide fragments. Bromelain, a plant protease derived from the stem of the pineapple plant, can even produce detectable semm proteolysis after oral adrninistration (180). Oral therapy with bromelain significantly reduces bmising that stems from obstetrical manipulations (181). Bromelain—pancreatin combinations have been more effective in digestive insufficiency compared to either pancreatin or placebo (182,183). Bromelain may also enhance the activity of antibiotics, especially tetracycline, when adrninistered concurrently (184). 

However, recent investigations on the effect of the tissue matrix on the detection limits attained by this test have indicated that ceftiofur, sulfonamides, streptomycin, and some macrolide antibiotics cannot be detected in intact meat with the plates and the bacterial strains prescribed in the European four-plate test (81, 82). Two plates of this system were not found suitable for screening sulfamethazine or streptomycin at levels far above the MRL the third plate detected tetracyclines and -lactams up to the MRL levels whereas the fourth was sensitive to -lactams and some but not all macrolides. Detection, on the other hand, of the fluoroquinolones enrofloxacin and ciprofloxacin could only be made possible by an additional Escherichia coli plate not included in the four-plate test. 

Key to the successful production of generic antisera for the tetracycline antibiotics has been the synthesis of the hapten 4-hydrazino-4-dedimethylamino-tetracycline and its subsequent conjugation to protein (73). The immunochemical method developed from this sera could detect tetracycline, chlortetracycline, and oxytetracycline residues in meat and milk, with adequate sensitivity (19). 

A number of tests were also developed and further used routinely to detect other process-related impurities in clinical batches, such as a Limulus amoebocyte lysate test to quantify bacterial endotoxins (< 1 I.U./mg of BBG2Na) and a specific RP-HPLC assay to detect tetracycline (no trace detected) since this antibiotic was used for selection by drug pressure during the fermentation step. 

A commercially available RIA developed as a screen for tetracycline antibiotics in serum, urine, milk, and tissue of livestock has been adapted by Meyer and co-workers to analyze water samples. The interest in pharmaceutical compounds in the enviromnent is relatively new. Immunoassays for pharmaceutical compounds are commonly used in biological media where the concentrations are quite high. Therefore, in this RIA the lower limit of antibiotic detection had to be modified to enable quantification of antibiotic levels as low as one part per billion in water samples. 

The literature data present some examples of sensors for the detection of several antibiotics and antihypertensive and anti-inflammatory drugs currently used in urban areas. The detection of antibiotic residues in environmental samples through the use of various disposable electrochemical sensors has proved the capacity of these sensors for fast, low-cost, and sensitive detection. Tetracyclines, as an important class of antibiotics, were investigated on gold SPEs using cyclic voltammetry... 

On the other hand, the analgesic phenylbutazone, the antidepressant paroxetine, tetracycline antibiotics doxycycline and chlorotetracycline, the antibiotics tilmico-sin, danofloxacin, enoxacin, nifuroxazide, the (3-blockers betaxolol, carazolol, pindolol, the (3-agonist clenbuterol, barbiturates, the antihypertensive lisinopril, and the drug to treat cancer tamoxifen were never detected. 

The usual method of detecting chromatographed tetracycline antibiotics involves fuming the paper with ammonia vapor and observing the yellow fluorescence under UV light. As little as 0.2-0.5 pg can be visualized by this technique. 

Macrolide antibiotics (clarithromycin, dehydroerythromycin, etc.) and sulfonamides (sulfamethoxazole, sulfadimethoxine, sulfamethazine, and sulfathi-azole) are the most prevalent antibiotics found in the environment with levels around a few micrograms per liter, whereas fluoroquinolones, tetracyclines, and penicillins have been detected in fewer cases and usually at low concentrations (nanograms per liter) [3,20,23,72]. This result is not surprising, since penicillins are easily hydrolyzed and tetracyclines readily precipitate with cations such as calcium and are accumulated in sewage sludge or sediments. Several reviews have reported the environmental occurrence of different antibiotics in aquatic and soil compartments. Some of these data are detailed in Table 1. 

Immunochemical methods have been developed and placed on the market to analyze tetracycline residues (see Table 4). Thus, a qualitative EIA has been developed and used to analyze tetracyclines in honey samples with a detection level of 20 pg/kg-1 [96]. A microplate-based indirect ELISA has been developed to analyze tetracyclines using polyclonal antibodies. The assay could measure tetracycline in the range between 0.1 and 6 ng mL L Other tetracycline antibiotics such as chlortetracycline, rolitetracycline, or minocycline are also highly recognized in this assay [98]. Several immunoassay kits are commercially available for the analysis of tetracyclines although, to our knowledge, none of them... 

As occurred with the other antibiotics, commercial immunoassay formats, also available as kits for tetracyclines and penicillins such as the Parallux, the LacTek, or the Charm II, have also been placed on the market for the analysis of sulfonamides (see Table 4). Thus, the Parallux detects sulfamethazine and sulfadimethoxine in raw milk with a LOD of 10 pg L1. The Charm II detects almost all sulfonamides in honey and milk with a LOD in the range from 1 to 10 pg L, whereas LacTek is able to detect sulfamethazine. Moreover, the 5101SULlp and 5101SUDAlp tests reach LOD values for sulfamethazine and sulfadiazine of around 0.2 pg L 1 and they have been applied to the analysis of urine, milk, and plasma. These tests have proved to be efficient as a point of care for on-site applications on farms. Moreover, commercially available antibodies can be found from several sources such as Silver Lake Research, US Biological, Cortex Biochem. Inc., Accurate Chemical Scientific, Fitzgerald Industries International Inc., and Biotrend Chemikalien GmbH. 

Zhou, J. K., Gerhardt, G. C., Baranski, A., and Cassidy, R. (1999). Capillary electrophoresis of some tetracycline antibiotics coupled with reductive fast cyclic yoltammetric detection. J. Chromatogr. A 839, 193-201. 

The conventional bioassays based on methodology developed by FDA and expanded by FSIS use four extractant buffers, five test organisms, five growth media, two incubation temperatures, and penicillinase to detect, identify, and/or quantify antibiotics such as the penicillins, streptomycins, tetracyclines, neomycins, erythromycin, tylosin, etc. Bioassay laboratory results are used by FSIS to take regulatory action and by FDA to prosecute farmers with histories of improperly withdrawing antibiotics before marketing their herds or flocks. 

FSIS laboratories also use chemical techniques and instrumentation to identify select antibiotic residues. The tetracyclines of interest are identified by thin layer chromatography. Sulfonamides are detected and quantified by fluorescence thin lay chromatography and confirmed by gas chromatography/mass spectrometry. Amoxicillin and gentamycin are identified and/or quantified by high pressure liquid chromatography. Similar techniques are used to identify ionophores and other antimicrobials of interest. 

Other sources of PPCP contamination to groundwater can originate from farms, leaking septic tanks, and lagoons. For instance, Campagnolo et al. (2002) detected several types of antibiotics including macrolides, tetracycline, sulfonamides, and (3-lactams in groundwater samples collected from sites that were in proximity of a swine farm. 

---